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Background/Objectives: Ulcerative colitis (UC) is a chronic and easily recurrent inflammatory bowel disease. The gut microbiota and plasma metabolites play pivotal roles in the development and progression of UC. Therefore, therapeutic strategies targeting the intestinal flora or plasma metabolites offer promising avenues for the treatment of UC. Luteolin (Lut), originating from a variety of vegetables and fruits, has attracted attention for its potent anti-inflammatory properties and potential to modulate intestinal flora. Methods: The therapeutic efficacy of Lut was evaluated in an established dextran sodium sulfate (DSS)-induced colitis mice model. The clinical symptoms were analyzed, and biological samples were collected for microscopic examination and the evaluation of the epithelial barrier function, microbiome, and metabolomics. Results: The findings revealed that Lut administration at a dose of 25 mg/kg significantly ameliorated systemic UC symptoms in mice, effectively reduced the systemic inflammatory response, and significantly repaired colonic barrier function. Furthermore, Lut supplementation mitigated gut microbiota dysbiosis in a UC murine model, increasing the abundance of Muribaculaceae, Rikenella, and Prevotellaceae while decreasing Escherichia_Shigella and Bacteroides levels. These alterations in gut microbiota also influenced plasma metabolism, significantly increasing phosphatidylcholine (PC), 6′-Deamino- 6′-hydroxyneomycin C, and gamma-L-glutamyl-butyrosine B levels and decreasing Motapizone and Arachidoyl-Ethanolamide (AEA) levels. Conclusions: This study reveals that Lut supplementation modulates intestinal inflammation by restoring the gut microbiota community structure, thereby altering the synthesis of inflammation-related metabolites. Lut is a potential nutritional supplement with anti-inflammatory properties and offers a novel alternative for UC intervention and mitigation. In addition, further studies are needed to ascertain whether specific microbial communities or metabolites can mediate the recovery from UC. [ABSTRACT FROM AUTHOR]

Simple Summary: Yeast protein feeds have the potential to replace other protein source feeds. Accordingly, in this paper, we used the industrial and agricultural byproducts of glucose master liquor and corn steep liquor as substrates, and utilized Candida utilis for fermentation. Firstly, the medium components of yeast protein feed were optimized, and then the optimized yeast protein feed was fed to weaned Dongliao black piglets; the feeding effect was analyzed using 16S rDNA and metabolomics. We found that Candida utilis feed has no adverse effects on the growth performance of the pigs and that it improves blood lipids and other indicators. A total of 10 plasma metabolites were correlated with 27 genera of bacteria, as revealed by the results of the combined multi-omics analysis, which were mainly enriched in the pathways of primary bile acid metabolism, histidine metabolism, and tryptophan metabolism. The results show that Candida utilis feed can maintain the growth performance of the pigs and enhance their immunity and maintenance of gut flora homeostasis; this likely occurs through amino acid metabolism. In this experiment, glucose master liquor and corn steep liquor were used as carbon and nitrogen sources, and Candida utilis was used as a strain to ferment yeast feed. The OD value and number of yeast cells were used as response values to optimize the medium components of the yeast feed through a response surface methodology. The optimal medium components were a glucose master liquor concentration of 8.3%, a corn steep liquor concentration of 1.2%, and a KH2PO4 concentration of 0.14%. Under this condition of fermentation, the OD value was 0.670 and the number of yeast cells was 2.72 × 108/mL. Then, we fed Candida utilis feed to Dongliao black piglets, and the effects of the yeast feed on the piglets' growth performance, fecal microbiota, and plasma metabolic levels were investigated through 16S rDNA sequencing and metabolomics. In total, 120 black piglets with an average initial weight of 6.90 ± 1.28 kg were randomly divided into two groups. One group was fed the basic diet (the CON group), and the other was supplemented with 2.5% Candida utilis add to the basic diet (the 2.5% CU group). After a pre-feeding period, the formal experiments were performed for 21 days. The results showed that the addition of Candida utilis to the diet did not affect growth performance compared with the control group. Meanwhile, no significant differences were observed in the serum biochemical indices. However, piglets in the 2.5% CU group had a significantly altered fecal microbiota, with an increased abundance of Clostridium_sensu_stricto_1, Lactobacillus, and Muribaculaceae_unclassified. Regarding the plasma metabolome, the 12 differential metabolites detected were mainly enriched in the histidine, tryptophan, primary bile acid, and caffeine metabolic pathways. Regarding the integrated microbiome–metabolome analysis, differential metabolites correlated with fecal flora to variable degrees, but most of them were beneficial bacteria of Firmicutes. Collectively, dietary Candida utilis feed had no adverse effect on growth performance; however, it played an important role in regulating fecal flora and maintaining metabolic levels. [ABSTRACT FROM AUTHOR]

Children are disproportionately represented among those who suffer asthma, which is a kind of chronic airway inflammation. Asthma symptoms might worsen when exposed to the air pollutant particulate matter 2.5 (PM2.5). However, it is becoming more prevalent among older adults, with more asthma-related deaths occurring in this pollution than in any other age group, and symptoms caused by asthma can reduce the quality of life of the elderly, whose asthma is underdiagnosed due to physiological factors. Therefore, in an effort to discover a therapy for older asthma during exposure to air pollution, we sought to ascertain the effects of pre-exposure (PA) and persistent exposure (PAP) to PM2.5 in aged asthma rats. In this study, we exposed aged rats to PM2.5 at different times (PA and PAP) and established an ovalbumin-mediated allergic asthma model. The basic process of elderly asthma caused by PM2.5 exposure was investigated by lung function detection, enzyme-linked immunosorbent assay (ELISA), histopathology, cytology, cytokine microarray, untargeted metabolomics, and gut microbiota analysis. Our findings demonstrated that in the PA and PAP groups, exposure to PM2.5 reduced lung function and exacerbated lung tissue damage, with varying degrees of effect on immunoglobulin levels, the findings of a cytological analysis, cytokines, and chemokines. The PA and PAP rats had higher amounts of polycyclic aromatic hydrocarbons (PAHs), such as naphthalene, 2-methylNaphthalene, 1-methylNaphthalene and flourene. Moreover, exposure to PM2.5 at different times showed different effects on plasma metabolism and gut microbiota. Bioinformatics analysis showed a strong correlation between PAHs, cytokines, and gut microbiota, and PAHs may cause metabolic disorders through the gut microbiota. These findings point to a possible mechanism for the development of asthma in older people exposure to PM2.5 that may be related to past interactions between PAHs, cytokines, gut microbiota, and plasma metabolites.

Fine particulate matter 2.5 (PM 2.5 ) is a prevalent atmospheric pollutant that is closely associated with asthma. Elderly patients have a high incidence of asthma with a long course of illness. Our previous studies revealed that exposure to PM 2.5 diminishes lung function and exacerbates lung damage in elderly rats. In the present study, we investigated whether PM 2.5 exposure influences susceptibility to allergic asthma in elderly rats. Brown-Norway elderly rats were treated with ovalbumin (OVA) for different durations before and after PM 2.5 exposure. The results from pulmonary function tests and histopathology indicated that early exposure to allergens prior to PM 2.5 exposure increased susceptibility to airway hyperresponsiveness and led to severe lung injury in elderly asthmatic rats. Cytokine microarray analysis demonstrated that the majority of cytokines and chemokines were upregulated in OVA-treated rats before and after PM 2.5 exposure. Cytological examination showed no change in eosinophil (EOS) counts, yet the amounts of neutrophils (NEU), white blood cells (WBC), lymphocytes (LYM), and monocytes (MON) in the lung lavage fluid of OVA-treated rats were significantly higher than those in control rats before and after PM 2.5 exposure, suggesting that PM 2.5 affects noneosinophilic asthma in elderly rats. ELISA results from the plasma and lung lavage fluid revealed that the levels of IgG1, IgE, IgG2a and IgG2b were significantly elevated in OVA-treated rats, whereas the level of IgG2b in the lung lavage fluid was significantly lower in rats treated with OVA prior to PM 2.5 exposure compared to those treated afterward. A non-targeted metabolomic analysis of plasma identified 202 metabolites, among which 31 metabolites were differentially abundant. Ten metabolites and 11 metabolic pathways were uniquely detected in OVA-treated rats before PM 2.5 exposure. Specifically, there were positive or negative correlations between the levels of Th2-associated cytokines (IL-4, IL-5, and IL-13) and six metabolites in the OVA-treated group before PM 2.5 exposure, whereas the levels of IL-4 and IL-5 were negatively correlated with five metabolites in the OVA-treated group after PM 2.5 exposure. Our findings suggest that PM 2.5 exposure could influence the susceptibility of allergic asthma in response to allergens in elderly rats, potentially through changes in plasma metabolites., Competing Interests: Competing interests: The authors declare no competing interests. Ethics approval and consent to participate: The Institute of Laboratory Animal Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College’s Institutional Animal Care and Use Committee approved this study (approval no. QC 20012) and agreed to carry out relevant animal experiments. In the course of the experiment, to ensure the implementation of the experimental animal welfare, ethical measures were taken. The study is reported in accordance with ARRIVE guidelines., (© 2025. The Author(s).)

Preeclampsia (PE) is a challenge in maternal healthcare due to its complex nature, characterized by high blood pressure, protein in the urine, and damage to various organs. There is evidence linking PE to endothelial dysfunction (ED), triggered by substances released from an oxygen-deprived placenta. Previous in vitro studies have not considered the impact of in vivo elements, such as the different patterns of blood flow, and laminar (LSS) vs. oscillatory (OSS) shear stress, on the development of ED. We investigated the impact of plasma from healthy pregnant women (HP), subjects with gestational hypertension (GH), and PE patients on global gene expression of human coronary endothelial cells (HCAECs) under LSS and OSS. Our findings revealed a unique transcriptional profile of endothelial cells induced by plasma incubation in LSS. Notably, OSS resulted in similar transcriptomes irrespective of plasma treatment. Under LSS, GH plasma resulted in a proliferative profile, whereas PE plasma was linked to pro-inflammatory and antioxidant profiles compared to HP plasma. Our findings demonstrate that shear stress levels influence the endothelial cell transcriptome in response to plasma from hypertensive pregnancy patients. Both PE and GH can induce endothelial dysfunction under atheroprotective LSS, with a more significant effect observed with PE-derived plasma., Competing Interests: Declarations. Competing interests: The authors have no interests to declare that are relevant to the content of this article., (© 2025. The Author(s).)

Plasma contains metabolites with diverse physicochemical properties, ranging from highly polar to highly apolar, and concentrations spanning at least nine orders of magnitude. Plasma metabolome analysis is valuable for monitoring health and evaluating medical interventions but is challenging due to the metabolome's diversity and complexity. This study aims to develop and validate targeted LC-MS/MS methods for quantifying 235 mammalian metabolites from 17 compound classes in porcine plasma without prior derivatization. Utilizing reversed-phase and hydrophilic interaction liquid chromatography coupled with tandem mass spectrometry, each analyte is identified and quantified using two selected reaction monitoring (SRM) transitions. Fast polarity switching and scheduled SRM enhance the metabolome coverage and throughput, enabling the analysis of one sample in about 40 min. A simple "dilute and shoot" sample preparation protocol was employed, with samples injected at two dilution levels to align metabolite concentrations within calibration curve ranges. Validation in porcine plasma included assessments of carryover, linearity, detection and quantification limits, repeatability and recovery. The method was further applied to plasma samples from various animal species, demonstrating its applicability to human and animal studies. This study establishes two robust LC-MS/MS methods for comprehensive porcine plasma metabolome quantification, advancing large-scale targeted metabolomics in biomedical research.

Birth asphyxia can result in kidney dysfunction, disturbances in systemic electrolytes and fluid balance in newborns. Currently, there is no proven dietary approach to support asphyxiated newborns. This study investigates whether oral plasma supplementation improves kidney function and overall health in asphyxiated newborns. Cesarean-delivered near-term pigs with or without an 8 min intrauterine clamping of the umbilical cord were fed a milk replacer dissolved in water for 24 h in Experiment 1. Pigs were fed 72 h with milk replacers dissolved in either maternal plasma or water in Experiment 2. Blood, urine, and kidney tissue were collected for further analyses. Asphyxia disrupted blood electrolyte balance. And plasma feeding led to higher fluid retention for both asphyxiated and control pigs. Additionally, plasma feeding may also affect kidney development and protect kidneys from asphyxia induced impairments. Birth asphyxia in pigs led to immediate disturbance of electrolyte balance, impaired fluid retention, and kidney impairments. Plasma feeding may improve postnatal newborn hydration and may also improve the condition of kidneys following asphyxia., (© 2025 The Author(s). Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

Vanillic acid (VA) is a dietary benzoic acid derivative, flavoring agent, and food stabilizer. In this study, the antioxidant and anti-inflammatory potential of VA was explored in vitro and ex vivo in human immune cells and non-cellular models. In neutrophils, VA significantly downregulated the f MLP-induced oxidative burst and the generation of reactive oxygen species (ROS); it also suppressed the release of pro-inflammatory cytokines (TNF-α, IL-8) and the tissue-remodeling enzyme elastase-2 (ELA-2) in cells stimulated with LPS and f MLP+cytochalasin B. Additionally, VA showed good biocompatibility with human neutrophils and peripheral blood mononuclear cells (PBMCs) across the tested concentrations of 1-50 µg/mL. Furthermore, VA at 1-5 μg/mL enhanced the non-enzymatic antioxidant capacity of human plasma (NEAC) and prevented oxidative and nitrative damage to plasma proteins by protecting tyrosine moieties and thiols from peroxynitrite. VA also inhibited lipid peroxidation and the formation of thiobarbituric acid-reactive substances (at 50 μg/mL) and protein-bound carbonyls (at 5-50 μg/mL) in peroxynitrite-treated plasma. In non-cellular tests, VA acted as a hypochlorous acid and hydrogen peroxide scavenger and inhibited non-enzymatic protein glycation, outperforming the references Trolox and aminoguanidine. Along with existing data from animal models and studies on polyphenol intake, these results might support the synergic role of VA in dietary protection against chronic diseases related to oxidative stress and inflammation.

Mass spectrometry-based proteomics is widely applied to human blood serum or plasma in the search of biomarkers for various diseases. However, the enormous complexity and dynamic range of protein concentrations in these samples render a significant analytical challenge, particularly for detecting low-abundance candidate biomarkers. As a result, strategies for enriching low-abundance proteins and improving their identification in serum or plasma proteomics are commonly used. Here, we describe an immunodepletion technique that is routinely used in our lab for removing high-abundance proteins from human serum/plasma., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Extracellular vesicles (EVs) are nanosized particles that are released by various cell types and play vital roles in intercellular communication. They carry biological molecules reflecting the physiological and pathological states of their source cells and tissues, showing potential as biomarkers. However, the impact of demographic factors like age and sex on the properties of blood plasma EVs remains underexplored. This study aims to fill this gap by evaluating how these factors influence the particle count and proteomic profiles of plasma EV preparations and corresponding protein fractions. Plasma samples from 120 healthy volunteers were collected and pooled into six groups: young males (age: 27.6 ± 4.0), young females (27.4 ± 3.8), middle-aged males (48.8 ± 3.8), middle-aged females (48.9 ± 3.9), old males (69.3 ± 3.9), and old females (69.4 ± 4.3). EV- and protein-enriched fractions were separated by size-exclusion chromatography (SEC). Fractions were characterized for particle number concentration and protein composition to identify characteristics affected by age and biological sex. Plasma EVs and corresponding protein fractions exhibited distinct characteristics, with differential enrichment of markers related to EVs and other blood components, including lipoproteins. Proteomic profiles of both EVs and protein fractions displayed sex- and age-dependent differences. Differentially abundant proteins displayed functions previously identified in the context of aging and sex differences, highlighting their utility as biomarkers. Age and sex significantly affect the characteristics of plasma EVs and proteins, potentially influencing their efficacy and interpretation as biomarkers in clinical applications. This study lays the groundwork for detailed mechanistic research to understand how EVs mediate age- and sex-related effects in health., (© 2024 The Author(s). Aging Cell published by Anatomical Society and John Wiley & Sons Ltd.)

Capillary electrophoresis coupled to mass spectrometry (CE-MS) has emerged as a powerful analytical technique with significant implications for clinical research and diagnostics. The integration of information from CE and MS strengthens confidence in the identification of compounds present in clinical samples. The ability of CE to separate molecules based on their electrophoretic mobility coupled to MS enables the accurate identification and quantification of analytes, even in complex biological matrices such as human plasma.Here, we present a detailed protocol for an untargeted metabolomics study using CE-MS and its application in a study on human plasma from patients suffering Long COVID syndrome. The protocol ranges from sample preparation to biological interpretation, detailing a workflow enabling the analysis of cationic and anionic compounds, metabolite identification, and data processing., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Hemolysis is the most prevalent pre-analytical interfering factor and a major source of error in laboratory analysis. The examination of samples post-centrifugation can provide valuable information regarding pre-analytical interferences. In this unusual case, a patient's plasma specimen was cherry-red after centrifugation, which is most usually indicative of hemolysis. However, subsequent investigations ruled out common hemolysis causes. We eventually determined that the patient's cherry-red plasma was most likely caused by other factors in the patient's medical history, including cancer treatment with PV-10 (rose bengal disodium 10%). We then conducted an interference study to comprehensively assess the effects of PV-10 on various biochemical tests, especially liver function tests and bilirubin levels. The findings indicate that PV-10 has varying effects on different biochemical assays and test results should be examined individually. This report underlines the need for awareness of potential drug interference on laboratory tests for better result interpretation and making clinical decisions., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Extracellular nanoparticles (EPs) are a subject of increasing interest for their biological role as mediators in cell-cell communication; however, their harvesting and assessment from bodily fluids are challenging, as processing can significantly affect samples. With the aim of minimizing processing artifacts, we assessed the number density ( n ) and hydrodynamic diameter ( D h ) of EPs directly in diluted plasma and blood using the following recently developed technique: interferometric light microscopy (ILM). We analyzed 613 blood and plasma samples from human patients with inflammatory bowel disease (IBD), collected in trisodium citrate and ethylenediaminetetraacetic acid (EDTA) anticoagulants, and 163 blood and plasma samples from canine patients with brachycephalic obstructive airway syndrome (BOAS). We found a highly statistically significant correlation between n in the plasma and n in the blood only in the human (i.e., but not canine) blood samples, between the samples with trisodium citrate and EDTA, and between the respective D h for both species (all p < 10 -3 ). In the human plasma, the average < D h > was 139 ± 31 nm; in the human blood, < D h > was 158 ± 11 nm; in the canine plasma, < D h > was 155 ± 32 nm; and in the canine blood, < D h > was 171 ± 33 nm. The differences within species were statistically significant ( p < 10 -2 ), with sufficient statistical power (P > 0.8). For < n >, we found no statistically significant differences between the human plasma and blood samples or between the samples with trisodium citrate and EDTA. Our results prove that measuring n and D h of EPs in minimally processed fresh blood and in diluted fresh plasma by means of ILM is feasible for large populations of samples.

Objectives: In most countries the majority of patients are in outpatient care. In difference to hospitalized patients, their blood samples often take hours after collection to centrifugation. The study investigates the release of potassium and the development of pseudohyperkalemia in lithium heparin (Li-Hep) and serum blood collection tubes over time., Methods: From 201 donors 4 serum and 4 Li-Hep blood collection tubes were taken each. After 0.5, 4, 6 and 8h whole blood was centrifuged, and potassium levels were determined. To simulate the preanalytic conditions, the samples with a storage time >0.5h were shaken on a standard shaker for 1h and stored at 4-8°C for the remaining time., Results: Over time, significant more potassium was released before centrifugation from the Li-Hep plasma than from serum (1.21 vs 0.94 mmol/L). After 6h, the two groups were no longer highly statistically significantly different (potassium mean: 5.01 mmol/L in serum group, 4.92 mmol/L in Li-Hep group). In the Li-Hep group 164 donors developed a pseudohyperkalemia after 8h, compared to 76 in the serum group., Significance: The decision as to which material is best suited should not only be based on which value comes closest to the physiological situation immediately after blood collection. The subsequent preanalytic circumstances must also be considered. Serum tubes appear to be at least as suitable for potassium determination as Li-Hep tubes. In terms of patient blood management, serum provides the possibility of performing a wider range of analyses in the outpatient setting., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Reuter et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The effectiveness of indirect Intravascular laser irradiation of blood (ILIB) is not fully understood. In this study, we provided a novel experiment that employs metabolomics to investigate the effects of ILIB in women. Twenty-eight volunteers underwent indirect ILIB and had their plasma collected before and after this procedure. The ILIB was applied at the radial artery for 30 min, using low-power photobiomodulation (660 nm), and a power output of 0.1 W. Plasma samples were extracted and analyzed using liquid chromatography-high-resolution mass spectrometry in an untargeted approach. Partial Least Squares Discriminant Analysis revealed 151 molecules with the Variable In Projection score of ≥ 1. From these, 26 were identified. After checking for molecules related to dietary intake, fasting, medication, or part of the human exposome, 15 were affected by ILIB. The abundances of Estradiol 17b-glucuronide 3-sulfate, CAR 14:3, PI 22:6/PGJ2, and CAR 12:1 significantly increased by ILIB, while AcylGlcADG 62:9, Tyrosyl-Glutamine, and CDP-DG 22:3/PGF1 had the contrary effect. ILIB was shown to modulate molecules from different chemical classes, although its impact on plasma metaboloma was minimal. Further research is warranted to fully elucidate the implications of these findings across various metabolic pathways, thus advancing the science surrounding ILIB., Competing Interests: Declarations. Competing interests: The authors declare no competing interests. Ethical approval: This study was conducted by the Helsinki criteria and approved by the Ethics Committee of São Francisco University (Approval Number: 63939422.2.0000.5514). Patient consent: All patients consented to participate in the study, providing their approval for this purpose., (© 2024. The Author(s).)

This study addresses the valorization of duck blood, an underutilized protein-rich by-product from the poultry industry, into bioactive protein hydrolysates with antioxidant and ACE inhibitory properties. Raw and heat-treated duck blood were compared as substrates for enzymatic hydrolysis using Neutrase and Papain. Gel electrophoresis revealed that heat treatment reduced fibrinogen content, while FTIR analysis showed that heat treatment modified the protein structure, increasing β-sheet content from 21.13 % to 34.96 %. Heat-treated duck blood hydrolyzed by Neutrase exhibited superior hydrolysis (9.53 %) and protein recovery (60.28 %) compared to raw blood. Pilot-scale production (1000 L) enhanced hydrolysate yield and maintained bioactive properties. LC-MS/MS analysis identified five novel bioactive peptides derived from the hydrolysate's 4-h simulated gastrointestinal (GI) digestion, with WMHVR demonstrating the highest antioxidant and ACE inhibitory activities. Molecular docking simulations revealed that WMHVR competitively inhibits ACE by binding to the S1, S2, and S' pockets through van der Waals and hydrogen bonding interactions. The GI-hydrolysate and identified peptides maintained bioactivity during simulated GI and blood plasma digestion, with ACE inhibition increasing over time. This research transforms food industry waste into functional protein hydrolysates, offering applications in nutraceuticals and functional foods while promoting sustainable practices through waste protein valorization., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Papungkorn Sangsawad reports financial support was provided by Agricultural Research Funding Agency of Thailand (Public Organization) (grant no. CRP6405031940). Papungkorn Sangsawad reports financial support was provided by Thailand Science Research and Innovation (TSRI), and the National Science, Research, and Innovation Fund (NSRF) (project codes: FF3-303-66-24-11(B)). Papungkorn Sangsawad reports financial support was provided by Full-time Doctoral Researcher (project codes: FtR.66/25/2567). If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Background: The use of whole blood compared with a balanced ratio of components in trauma resuscitation remains an area of ongoing investigation. One factor that may affect outcomes is the age of the blood product transfused. We used a murine model of blood banking and hemorrhagic shock resuscitation to compare the effect of storage duration in whole blood and packed red blood cells on the recipient inflammatory response., Methods: Murine whole blood or packed red blood cells were evaluated for the red blood cells storage lesion up to 14 days. Mice underwent hemorrhagic shock followed by resuscitation with whole blood or packed red blood cells combined with equal volume of thawed plasma (1:1) stored for 1, 7, or 14 days. Serum and lung cytokine/chemokine levels were measured and leukocyte infiltration determined via immunohistochemistry., Results: Both whole blood and packed red blood cells develop a blood storage lesion. Four hours after resuscitation, mice resuscitated with either day 14 whole blood or 1:1 demonstrated increased inflammatory cytokines and chemokines with similar findings within lung tissue compared with mice resuscitated with whole blood and 1:1 products stored for 1 or 7 days., Conclusions: Resuscitation with murine packed red blood cells or whole blood stored for 14 days produces a pronounced recipient inflammatory response compared with those units stored for lesser durations. Given the shorter storage duration of human whole blood to packed RBCs, resuscitation with whole blood within current storage limits may represent an advantageous resuscitation strategy compared with older packed red blood cells., Competing Interests: Conflict of Interest/Disclosure Dr Goodman reports receiving honoraria from Johnson & Johnson and Grifols and serving as expert testimony for Poling Law. Dr Pritts reports receiving honoraria from the World Trauma Symposium of the National Association of Emergency Medical Technicians. Dr Sisak, Dr Caldwell, Mr. Nguyen, Mr. Joseph, Ms. Schuster, and Ms. England report no biomedical financial interests or potential conflicts of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: Plasma from patients with active thyroid-associated orbitopathy (TAO-A) could cause inflammation to fibroblasts, and such a mechanism was explored in the context of melanoma., Methods: Plasma samples collected from TAO-A patients and healthy control (HC) were primarily co-cultured with the melanoma-associated fibroblasts (MAFs) derived from melanoma patients. The survival and inflammation of the co-cultured MAFs were measured after confirming the levels of pro-inflammatory cytokines. Ki67 and Vimentin (VIM) markers were analyzed by immunofluorescence, and cell survival and migration were assessed using cell counting kit-8 (CCK-8) and Transwell. The THP-1 cells were induced to differentiate into macrophages, which were subsequently co-cultured to assess M1/M2 polarization status. Meanwhile, the levels of inflammatory factor were detected by enzyme-linked immunosorbent assay (ELISA). The gene expression was measured by reverse transcription quantitative PCR (RT-qPCR), and the activation of PI3K/AKT, STAT1, p65, and ERK signaling pathways was detected by Western Blot., Results: Plasmas derived from TAO-A patients were characterized by elevated levels of pro-inflammatory cytokines, which enhanced the inflammation status and survival of MAFs, promoted the levels of PI3K and AKT, and downregulated expression of Bax. The co-culture of the plasma with MAFs evidently promoted M1 polarization and the phosphorylation of STAT1, P65 and ERK1/2., Conclusion: These findings proved the effects of the plasmas of TAO-A patients on the survival and inflammation of MAFs, providing evidence for future studies to delve into the relevant mechanisms., Competing Interests: The authors declare that they have no competing interests., (© 2024 Chen et al.)

Albumin is a major protein in mammalian blood plasma or serum, where its concentration in healthy organisms is about 600 μM [...].

The protein corona formed on nanoparticles (NPs) has potential as a valuable diagnostic tool for improving plasma proteome coverage. Here, we show that spiking small molecules, including metabolites, lipids, vitamins, and nutrients into plasma can induce diverse protein corona patterns on otherwise identical NPs, significantly enhancing the depth of plasma proteome profiling. The protein coronas on polystyrene NPs when exposed to plasma treated with an array of small molecules allows for the detection of 1793 proteins marking an 8.25-fold increase in the number of quantified proteins compared to plasma alone (218 proteins) and a 2.63-fold increase relative to the untreated protein corona (681 proteins). Furthermore, we discovered that adding 1000 µg/ml phosphatidylcholine could singularly enable the detection of 897 proteins. At this specific concentration, phosphatidylcholine selectively depletes the four most abundant plasma proteins, including albumin, thus reducing the dynamic range of plasma proteome and enabling the detection of proteins with lower abundance. Employing an optimized data-independent acquisition approach, the inclusion of phosphatidylcholine leads to the detection of 1436 proteins in a single plasma sample. Our molecular dynamics results reveal that phosphatidylcholine interacts with albumin via hydrophobic interactions, H-bonds, and water bridges. The addition of phosphatidylcholine also enables the detection of 337 additional proteoforms compared to untreated protein corona using a top-down proteomics approach. Given the critical role of plasma proteomics in biomarker discovery and disease monitoring, we anticipate the widespread adoption of this methodology for the identification and clinical translation of biomarkers., (© 2024. The Author(s).)

Extracellular vesicles (EVs) are a heterogeneous collection of particles that play a crucial role in cell-to-cell communication, primarily due to their ability to transport molecules, such as proteins. Thus, profiling EV-associated proteins offers insight into their biological effects. EVs can be isolated from various biological fluids, including donor blood components such as cryoprecipitate and fresh frozen plasma (FFP). In this study, we conducted a proteomic analysis of five single donor units of cryoprecipitate, FFP, and EVs derived from these blood components using a quantitative mass spectrometry approach. EVs were successfully isolated from both cryoprecipitate and FFP based on community guidelines. We identified and quantified approximately 360 proteins across all sample groups. Principal component analysis and heatmaps revealed that both cryoprecipitate and FFP are similar. Similarly, EVs derived from cryoprecipitate and FFP are comparable. However, they differ between the originating fluids and their derived EVs. Using the R-package MS-DAP, differentially expressed proteins (DEPs) were identified. The DEPs for all comparisons, when submitted for gene enrichment analysis, are involved in the complement and coagulation pathways. The protein profile generated from this study will have important clinical implications in increasing our knowledge of the proteins that are associated with EVs derived from blood components.

In cold human blood, the anomalous dynamics of adenosine triphosphate (ATP) result in the progressive accumulation of adenosine diphosphate (ADP), adenosine monophosphate (AMP), inosine monophosphate (IMP), inosine, and hypoxanthine. While the ATP, ADP, AMP, and IMP are confined to red blood cells (RBCs), inosine and hypoxanthine are excreted into plasma/serum. The plasma/serum levels of inosine and hypoxanthine depend on the temperature of blood and the plasma/serum contact time with the RBCs, and hence they represent robust biomarkers for evaluating the preanalytical quality of plasma/serum. These biomarkers are highly specific since they are generally absent or at very low levels in fresh plasma/serum and are highly sensitive since they are derived from ATP, one of the most abundant metabolites in blood. Further, whether blood was kept at room temperature or on ice could be predicted based on inosine levels. An analysis of >2000 plasma/serum samples processed for metabolomics-centric analyses showed alarmingly high levels of inosine and hypoxanthine. The results highlight the gravity of sample quality challenges with high risk of grossly inaccurate measurements and incorrect study outcomes. The discovery of these robust biomarkers provides new ways to address the longstanding and underappreciated preanalytical sample quality challenges in the blood metabolomics field.

This study aimed to determine the effects of spray dried plasma (SDP) on growth performance, carcass traits, tibia quality, and hemagglutination inhibition titers in broilers fed two nutritional strategies with high or low nutrient density. In the study, 816 one-day-old Ross 308 male broiler chickens were divided into a 2 × 2 factorial arrangements consisting of four treatment groups with 12 replicates (17 birds/replicate) based on diets with high nutrient density (HND) or low nutrient density (LND) from d 0 to 42 and receiving either control or 1% SDP diets during d 0 to 10. The results showed that feed intake (FI) and body weight gain (BWG) were increased (P < 0.05) and feed conversion ratio (FCR) was significantly reduced (P = 0.003) for broilers fed HND diets from d 0 to 42. The inclusion of SDP increased the BWG (P < 0.001), FI (P < 0.001), and FCR (P < 0.05) during d 0 to 10 of broiler life but not effect of SDP was observed for the whole 0-42 d period. Carcass yield increased with HND (P < 0.001) and dietary SDP (P = 0.002). However, HND feeding significantly decreased liver (P < 0.001), bursa of Fabricius (P = 0.002), abdominal fat (P < 0.001), proventriculus (P < 0.001) and gizzard weight (P < 0.001), but increased heart weight (P = 0.013), although spleen weight remained unaffected (P > 0.05) on d 42. Tibial bone morphometric and mechanical properties improved (P < 0.05) with SDP supplementation, and bone ash, Ca, and P remained unaffected (P > 0.05) on d 14. With the exception at d 28 (P = 0.037), the antibody titer to ND virus was similar among all treatment groups (P > 0.05) at d 0, 14, and 42. In conclusion, HND diets improve performance of broilers during the whole period and SDP supplementation during starter phase improve performance at this period, but also increased carcass yield, and tibial quality. Therefore, inclusion of SDP in the starter diet could be a beneficial nutritional strategy to improve the health and production of broilers provided feeding strategies using various nutrient densities., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Bundur et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

As climate change alters the hydric regime of many habitats, understanding the hydric physiology of animals becomes increasingly important. Plasma osmolality is a popular metric to assess an organism's hydration, but samples often need to be stored before being analyzed, under varying conditions and for different lengths of time. Previous studies on plasma storage conditions, and how they impact sample integrity, are minimal and have focused more on clinical applications than field studies. We studied the stability of osmolality values from wild rattlesnake plasma samples stored in commonly used plastic snap-cap tubes under different time (0, 2, 3, 7, 29 days) and temperature (refrigerated at 2 °C and frozen at -18 °C) treatments. We hypothesized that frozen samples would remain more stable (e.g., retain osmolality values more similar to baseline values) than refrigerated samples because freezing the plasma would reduce evaporation. We found that osmolality of samples increased over time at both temperatures, becoming significantly higher than baseline after 7 days. Contrary to our prediction, osmolality increased more in frozen samples than in refrigerated samples. We discuss possible reasons for our results, along with their implications. To obtain the most accurate plasma osmolality values, we recommend refrigerating plasma samples for as short a time as possible, 3 days or fewer, before analyzing them on an osmometer., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Metabolomics is an emerging and powerful bioanalytical method supporting clinical investigations. Serum and plasma are commonly used without rational prioritization. Serum is collected after blood coagulation, a complex biochemical process involving active platelet metabolism. This may affect the metabolome and increase the variance, as platelet counts and function may vary substantially in individuals. A multiomics approach systematically investigating the suitability of serum and plasma for clinical studies demonstrated that metabolites correlated well ( n = 461, R 2 = 0.991), whereas lipid mediators ( n = 83, R 2 = 0.906) and proteins ( n = 322, R 2 = 0.860) differed substantially between specimen. Independently, analysis of platelet releasates identified most biomolecules significantly enriched in serum compared to plasma. A prospective, randomized, controlled parallel group metabolomics trial with acetylsalicylic acid administered for 7 days demonstrated that the apparent drug effects significantly differ depending on the analyzed specimen. Only serum analyses of healthy individuals suggested a significant downregulation of TXB2 and 12-HETE, which were specifically formed during coagulation in vitro . Plasma analyses reliably identified acetylsalicylic acid effects on metabolites and lipids occurring in vivo such as an increase in serotonin, 15-deoxy-PGJ2 and sphingosine-1-phosphate and a decrease in polyunsaturated fatty acids. The present data suggest that plasma should be preferred above serum for clinical metabolomics studies as the serum metabolome may be substantially confounded by platelets.

Abnormal plasma uric acid (UA) levels, the lipid profile, and plasma proteins in blood are associated with a range of adverse health outcomes. This multicenter, prospective cohort study aimed to determine the possible effects of multiple apheresis plasma donations on plasma UA levels, the lipid profile, and major proteins in plasma donors. Participants were enrolled from 1 April 2021 to 31 August 2022. When their plasma UA (men: >420 µmol/L, women: >360 µmol/L) and/or lipid levels (total cholesterol [TC]: ≥6.2 mmol/L, triglycerides [TGs]: ≥2.3 mmol/L, low-density lipoprotein cholesterol: ≥4.1 mmol/L, or high-density lipoprotein cholesterol [HDL-C]: <1.0 mmol/L) were abnormal at their first plasma donation, the enrolled participants were followed up until they had completed 10 plasma donations. A total of 11485 participants were enrolled, of whom 1861 met the inclusion criteria. During the study period, 320 donors completed 10 plasma donations. None of the participants took any corrective medicine for their abnormal index. The measured parameters were significantly different from the first to the tenth plasma donations (donors with asymptomatic hyperuricemia: UA, P < 0.001; donors with asymptomatic hyperlipidemia: HDL-C, P < 0.001; TC, P = 0.025; TGs, P < 0.001; apolipoprotein B, P = 0.025; all of the plasma donors, immunoglobulin G, P < 0.001). The levels of HDL-C, TC, and apolipoprotein B were increased, and the levels of UA, TGs, and immunoglobulin G were decreased over this time. However, immunoglobulin G levels were still in the normal range. Moreover, the changes in these parameters were closely associated with the frequency of plasma donation during the study period. Repeated apheresis plasma donations can reduce plasma UA and TG levels and increase HDL-C levels; and further evaluation of the clinical significance with a larger sample size is required., Competing Interests: Conflict of interest statement The authors declare no conflict of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Blood analysis is an important tool for monitoring the health status of fish, but the time between collection and analysis can affect the outcome of the analysis. This study sought to determine the maximum time refrigerated blood and frozen plasma samples of the tambaqui, Colossoma macropomum, can be stored without affecting analysis. Samples from 12 fish were obtained, stored under refrigeration at 4 °C and evaluated after 0, 24, 48, 72, and 96 h, while samples from 14 fish were centrifuged, and the resulting plasma was frozen at -20 °C and then evaluated after 0, 8, 12, 16 and 20 weeks. The parameters analyzed were hematocrit (Ht), hemoglobin content (Hb), total erythrocytes (RBC), total (WBC) and differential leukocytes, total thrombocytes (TC), glucose content (Glc), total protein (TP), triglyceride content (TG), total cholesterol (CoT), and the activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST). For refrigerated whole blood samples, mean corpuscular hemoglobin content (MCHC) showed a transient decline in 24 h, and there were decreases in WBC, TC, Glc and TG that persisted until the 72 h sample point (for Glc and TG) or persisted until the 96 h sample point (for WBC and TC). A decrease in RBC was noted from 48 h on, while ALT was significantly higher in the 96 h sample. Significant decreases in lymphocytes, monocytes, neutrophils and eosinophils were noted from 48 h of storage on, while a significant decline in basophil counts were noted over the last two sampled timepoints. The coefficient of variation was greatest at the 96 h timepoint, indicating increased variability in measured parameters after 4 d of refrigeration. Plasma samples frozen at -20 °C showed a significant variation in ALT after 8 weeks, and increases in TP and TG after 20 weeks. Therefore, it is recommended that refrigerated tambaqui whole blood samples be analyzed within 24 h and frozen tambaqui plasma samples analyzed within 8 weeks., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The endothelial glycocalyx, a gel-like layer that lines the luminal surface of blood vessels, is composed of proteoglycans, glycoproteins, and glycosaminoglycans. The endothelial glycocalyx plays an essential role in vascular homeostasis, and its degradation in trauma and sepsis can lead to microvascular dysfunction and organ injury. While there are no proven therapies for preventing or treating endothelial glycocalyx degradation, some initial literature suggests that plasma may have a therapeutic role in trauma and sepsis patients. Overall, the literature suggesting the use of plasma as a therapy for endothelial glycocalyx degradation is non-clinical basic science or exploratory. Plasma is an established therapy in the resuscitation of patients with hemorrhage for restoration of coagulation factors. However, plasma also contains other bioactive components, including sphingosine-1 phosphate, antithrombin, and adiponectin, which may protect and restore the endothelial glycocalyx, thereby helping to maintain or restore vascular homeostasis. This narrative review begins by describing the endothelial glycocalyx in health and disease: we discuss the overlapping disease mechanisms in trauma and sepsis that lead to its damage and introduce plasma transfusion as a potential therapy for prevention and treatment of endothelial glycocalyx degradation. Second, we review the literature on plasma as an exploratory therapy for endothelial glycocalyx degradation in trauma and sepsis. Third, we discuss the safety of plasma transfusion by reviewing the adverse events associated with plasma and other blood product transfusions, and we examine modern transfusion precautions that have enhanced the safety of plasma transfusion. We conclude that the literature proposes that plasma may have the potential to prevent and treat endothelial glycocalyx degradation in trauma and sepsis, indicating the need for further research., (© 2024. The Author(s).)

Media supplemented with sera from acutely exercised men has been shown to have 'anti-cancer' effects on prostate and breast cancer cell lines. This study investigated whether media supplemented with plasma samples taken at rest (≥30 h since the most recent exercise session) from men who were endurance-trained (END), strength-trained (STR) or recreationally active controls (CON) impacted the results of four assays that mimic hallmarks of cancer (proliferation, migration, extracellular matrix invasion and anoikis resistance) in the BT-549 breast cancer cell line. Compared to control conditions of either serum-free media or fetal bovine serum as appropriate, BT-549 cells cultured with plasma-supplemented media regardless of group resulted in greater cell proliferation (∼20-50%) and cell migration (∼15-20%), and lower extracellular matrix invasion (∼10-20%) and anoikis resistance (∼15-20%). Supplementing media with plasma from END or STR did not impact any outcomes of these assays compared to plasma from CON. Media supplemented with human plasma can impact functional assays reflective of cancer hallmarks in BT-549 cells, but effects of exercise on proliferation, migration, extracellular matrix invasion and anoikis resistance were not evident in resting blood samples of individuals with a prior history of exercise training., (© 2023 The Authors. Experimental Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Here, we present a workflow for analyzing multi-omics data of plasma samples in patients with post-COVID condition (PCC). Applicable to various diseases, we outline steps for data preprocessing and integrating diverse assay datasets. Then, we detail statistical analysis to unveil plasma profile changes and identify biomarker-clinical variable associations. The last two steps discuss machine learning techniques for unsupervised clustering of patients based on their inherent molecular similarities and feature selection to identify predictive biomarkers. For complete details on the use and execution of this protocol, please refer to Wang et al. 1 ., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Red blood cells (RBCs) constitute ∼50% of the bloodstream and represent an important target for environmental pollutants and bacterial/viral infections, which can result in their rupture. In addition, diseases such as sickle cell anaemia and paroxysmal nocturnal haemoglobinuria can also result in the rupture of RBCs, which can be potentially life-threatening. With regard to the release of cytosolic metalloproteins from RBCs into the blood-organ system, the biochemical fate of haemoglobin is rather well understood, while comparatively little is known about another highly abundant Zn-metalloprotein, carbonic anhydrase (CA I). To gain insight into the interaction of CA I with human blood plasma constituents, we have employed a metallomics tool comprised of size-exclusion chromatography (SEC) coupled online with an inductively coupled plasma atomic emission spectrometer (ICP-AES), which allows to simultaneously observe all Cu, Fe, and Zn-metalloproteins. After the addition of CA I to human blood plasma incubated at 37°C, the SEC-ICP-AES analysis using phosphate buffered saline (pH 7.4) after 5 min, 1 h, and 2 h revealed that CA I eluted after all endogenous Zn-metalloproteins in the 30 kDa range. Matrix-assisted laser desorption-time of flight mass spectrometry analysis of the collected Zn-peak confirmed that CA I eluted from the column intact. Our in vitro results suggest that CA I released from RBCs to plasma remains free and may be actively involved in health-relevant adverse processes that unfold at the bloodstream-endothelial interface, including atherosclerosis and vision loss., (© The Author(s) 2024. Published by Oxford University Press.)

Alzheimer's disease (AD) is a progressive neurodegenerative disorder, and moderate exercise holds promise in ameliorating the ongoing neurodegeneration and cognitive decline. Here, we investigated whether exercise-enriched blood plasm could yield a beneficial therapeutic effect on AD pathologies and cognitive decline in transgenic AD (P301S) mice. In this investigation, a cohort of 2-month-old C57BL/6 mice were granted continuous access to either a running wheel or a fixed wheel for 6 weeks. After that, their plasmas were extracted and subsequently injected intravenously into 4.5-month-old P301S mice biweekly over a 6-week period. A comprehensive methodology was then employed, integrating behavioral tests, pathology assessments, and biochemical analyses to unveil the potential anti-dementia implications of exercise-enriched blood plasma in P301S mice. Upon systemic administration, the findings revealed a noteworthy attenuation of hippocampus-dependent behavioral impairments in P301S mice. Conversely, blood plasma from sedentary counterparts exhibited no discernible impact. These effects were intricately associated with the mitigation of neuroinflammation, the augmentation of hippocampal adult neurogenesis, and a reduction of synaptic impairments following the administration of exercise-enriched blood plasma. These findings advance the proposition that administering exercise-enriched blood plasma may serve as an effective prophylactic measure against AD, opening avenues for further exploration and potential therapeutic interventions., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Abstract: Individuals living with sickle cell disease (SCD) experience severe recurrent acute and chronic pain. Challenges to gaining mechanistic insight into pathogenic SCD pain processes include differential gene expression and function of sensory neurons between humans and mice with SCD, and extremely limited availability of neuronal tissues from patients with SCD. Here, we used induced pluripotent stem cells (iPSCs), derived from patients with SCD, differentiated into sensory neurons (SCD iSNs) to begin to overcome these challenges. We characterize key gene expression and function of SCD iSNs to establish a model to investigate intrinsic and extrinsic factors that may contribute to SCD pain. Despite similarities in receptor gene expression, SCD iSNs show pronounced excitability using patch clamp electrophysiology. Furthermore, we find that plasma taken from patients with SCD during acute pain associated with a vaso-occlusive event increases the calcium responses to the nociceptive stimulus capsaicin in SCD iSNs compared with those treated with paired plasma from patients with SCD at steady state baseline or healthy control plasma samples. We identified high levels of the polyamine spermine in baseline and acute pain states of plasma from patients with SCD, which sensitizes SCD iSNs to subthreshold concentrations of capsaicin. Together, these data identify potential intrinsic mechanisms within SCD iSNs that may extend beyond a blood-based pathology., (© 2024 American Society of Hematology. Published by Elsevier Inc. All rights are reserved, including those for text and data mining, AI training, and similar technologies.)

In the present study, an enzymatically hydrolyzed porcine plasma (EHPP) was nutritionally and molecularly characterized. EHPP molecular characterization showed, in contrast to spray-dried plasma (SDP), many peptides with relative molecular masses (Mr) below 8,000, constituting 73% of the protein relative abundance. IIAPPER, a well-known bioactive peptide with anti-inflammatory and antioxidant properties, was identified. In vivo functionality of EHPP was tested in C. elegans and two different mouse models of intestinal inflammation. In C. elegans subjected to lipopolysaccharide exposure, EHPP displayed a substantial anti-inflammatory effect, enhancing survival and motility by 40% and 21.5%, respectively. Similarly, in mice challenged with Staphylococcus aureus enterotoxin B or Escherichia coli O42, EHPP and SDP supplementation (8%) increased body weight and average daily gain while reducing the percentage of regulatory Th lymphocytes. Furthermore, both products mitigated the increase of pro-inflammatory cytokines expression associated with these challenged mouse models. In contrast, some significant differences were observed in markers such as Il-6 and Tnf-α, suggesting that the products may present different action mechanisms. In conclusion, EHPP demonstrated similar beneficial health effects to SDP, potentially attributable to the immunomodulatory and antioxidant activity of its characteristic low Mr bioactive peptides., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Solà-Ginés et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

In this paper, we studied the potential genotoxic effects of human plasma from healthy volunteers, as well as patients with gastro-oesophageal reflux disease, Barrett's oesophagus (BO) and oesophageal adenocarcinoma (OAC) using the oesophageal adenocarcinoma cell line (OE33) and the lymphoblastoid cell line (TK6). Both TK6 and OE33 cells were treated with plasma (10 % volume, replacing foetal bovine serum (FBS) or horse serum (HS)) at different time points of 4 h (for the micronucleus (Mn) assay and the invasion assay) and 24 h (for the cell cycle studies). Plasma-induced effects on DNA damage levels, cell viability and the cell cycle were studied by the micronucleus assay, cytokinesis block proliferation index (CBPI) and flow cytometry respectively. The expression of IL-8 in supernatants of TK6 cells and IFN-β in OE33 cells was also analysed by enzyme-linked immunosorbent assay (ELISA). Finally, we carried out an assessment of cellular invasion of OE33 cells following plasma treatment. The results of the micronucleus assay confirmed the genotoxicity of direct plasma treatment from some participants through the increase in DNA damage in TK6 cells. Conversely, some individual patient plasma samples reduced background levels of TK6 cell Mn frequency, in an anti-genotoxic fashion. In TK6 cells, (on average) plasma samples from patients with Barrett's oesophagus induced higher micronucleus levels than healthy volunteers (p= 0.0019). There was little difference in Mn induction when using plasma versus serum to treat the cells in vitro. Cell cycle results showed that direct plasma treatment had a marked impact on OE33 cells at 24 h (p=0.0182 for BO and p=0.0320 for OAC) by decreasing the proportion of cells in the S phase, while plasma exposure was less impactful on the cell cycle of TK6 cells. Invasion of OE33 cells was also seen to be non-significantly affected by plasma treatment of OE33 cells. The addition of N-acetyl cysteine NAC in a dose-dependent matter did not alter the formation of Mn in TK6 cells, suggesting that reactive oxygen species (ROS) are not the root cause of plasma's genotoxicity. The concentration of IL-8 in TK6 cells and IFN-β in OE33 cells was significantly higher in cells treated with OAC-derived plasma than in the untreated negative control. Collectively, our results demonstrate that plasma-specific effects are detectable which helps us better understand some important aspects of the biology of blood-based biomarkers under development., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Objective: The aim of this study was to produce and characterize triple-layered cell sheet constructs with varying cell compositions combined or not with the fibrin membrane scaffold obtained by the technology of Plasma Rich in Growth Factors (mPRGF)., Materials and Methods: Human primary cultures of periodontal ligament stem cells (hPDLSCs) were isolated, and their stemness nature was evaluated. Three types of triple-layered composite constructs were generated, composed solely of hPDLSCs or combined with human umbilical vein endothelial cells (HUVECs), either as a sandwiched endothelial layer or as coculture sheets of both cell phenotypes. These three triple-layered constructs were also manufactured using mPRGF as cell sheets' support. Necrosis, glucose consumption, secretion of extracellular matrix proteins and synthesis of proangiogenic factors were determined. Histological evaluations and proteomic analyses were also performed., Results: The inclusion of HUVECs did not clearly improve the properties of the multilayered constructs and yet hindered their optimal conformation. The presence of mPRGF prevented the shrinkage of cell sheets, stimulated the metabolic activity and increased the matrix synthesis. At the proteome level, mPRGF conferred a dramatic advantage to the hPDLSC constructs in their ability to provide a suitable environment for tissue regeneration by inducing the expression of proteins necessary for bone morphogenesis and cellular proliferation., Conclusions: hPDLSCs' triple-layer construct onto mPRGF emerges as the optimal structure for its use in regenerative therapeutics., Clinical Relevance: These results suggest the suitability of mPRGF as a promising tool to support cell sheet formation by improving their handling and biological functions., Competing Interests: Competing interest The authors declare the following competing financial interests: E.A. is the Scientific Director of and M.T, M.Z., R.T., and M.H.A. are scientists at BTI Biotechnology Institute, a dental implant company that investigates in the fields of oral implantology and PRGF-Endoret technology. N.O., A.F. and A.M.Z. declare no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Introduction: Untargeted metabolomics studies are expected to cover a wide range of compound classes with high chemical diversity and complexity. Thus, optimizing (pre-)analytical parameters such as the analytical liquid chromatography (LC) column is crucial and the selection of the column depends primarily on the study purpose., Objectives: The current investigation aimed to compare six different analytical columns. First, by comparing the chromatographic resolution of selected compounds. Second, on the outcome of an untargeted toxicometabolomics study using pooled human liver microsomes (pHLM), rat plasma, and rat urine as matrices., Methods: Separation and analysis were performed using three different reversed-phase (Phenyl-Hexyl, BEH C 18 , and Gold C 18 ), two hydrophilic interaction chromatography (HILIC) (ammonium-sulfonic acid and sulfobetaine), and one porous graphitic carbon (PGC) columns coupled to high-resolution mass spectrometry (HRMS). Their impact was evaluated based on the column performance and the size of feature count, amongst others., Results: All three reversed-phase columns showed a similar performance, whereas the PGC column was superior to both HILIC columns at least for polar compounds. Comparing the size of feature count across all datasets, most features were detected using the Phenyl-Hexyl or sulfobetaine column. Considering the matrices, most significant features were detected in urine and pHLM after using the sulfobetaine and in plasma after using the ammonium-sulfonic acid column., Conclusion: The results underline that the outcome of this untargeted toxicometabolomic study LC-HRMS metabolomic study was highly influenced by the analytical column, with the Phenyl-Hexyl or sulfobetaine column being the most suitable. However, column selection may also depend on the investigated compounds as well as on the investigated matrix., (© 2024. The Author(s).)

Pregnancy at advanced maternal age (AMA) is a condition of potential risk for the development of maternal-fetal complications with possible repercussions even in the long term. Here, we analyzed the changes in plasma redox balance and the effects of plasma on human umbilical cord mesenchymal cells (hUMSCs) in AMA pregnant women (patients) at various timings of pregnancy. One hundred patients and twenty pregnant women younger than 40 years (controls) were recruited and evaluated at various timings during pregnancy until after delivery. Plasma samples were used to measure the thiobarbituric acid reactive substances (TBARS), glutathione and nitric oxide (NO). In addition, plasma was used to stimulate the hUMSCs, which were tested for cell viability, reactive oxygen species (ROS) and NO release. The obtained results showed that, throughout pregnancy until after delivery in patients, the levels of plasma glutathione and NO were lower than those of controls, while those of TBARS were higher. Moreover, plasma of patients reduced cell viability and NO release, and increased ROS release in hUMSCs. Our results highlighted alterations in the redox balance and the presence of potentially harmful circulating factors in plasma of patients. They could have clinical relevance for the prevention of complications related to AMA pregnancy.

Plasma metabolomics holds potential for precision medicine, but limited information is available to compare the performance of such methods across multiple cohorts. We compared plasma metabolite profiles after an overnight fast in 11,309 participants of five population-based Swedish cohorts (50-80 years, 52% women). Metabolite profiles were uniformly generated at a core laboratory (Metabolon Inc.) with untargeted liquid chromatography mass spectrometry and a comprehensive reference library. Analysis of a second sample obtained one year later was conducted in a subset. Of 1629 detected metabolites, 1074 (66%) were detected in all cohorts while only 10% were unique to one cohort, most of which were xenobiotics or uncharacterized. The major classes were lipids (28%), xenobiotics (22%), amino acids (14%), and uncharacterized (19%). The most abundant plasma metabolome components were the major dietary fatty acids and amino acids, glucose, lactate and creatinine. Most metabolites displayed a log-normal distribution. Temporal variability was generally similar to clinical chemistry analytes but more pronounced for xenobiotics. Extensive metabolite-metabolite correlations were observed but mainly restricted to within each class. Metabolites were broadly associated with clinical factors, particularly body mass index, sex and renal function. Collectively, our findings inform the conduct and interpretation of metabolite association and precision medicine studies., (© 2024. The Author(s).)

Thrombotic thrombocytopenic purpura (TTP) is a life-threatening thrombotic disorder associated with a severe deficiency of ADAMTS-13-the protease that cleaves von Willebrand factor. Plasma therapy is the current standard of care for managing acute episodes of TTP, which involves removing patient plasma and replacing it with donor plasma to raise the level of ADAMTS-13 activity. Recently, therapies aimed at replacing ADAMTS-13 have been investigated as possible substitutes or add-ons to plasma therapy for congenital and immune-mediated TTP. Enzyme replacement therapy provides recombinant ADAMTS-13 via intravenous (i.v.) infusion to restore enzyme activity. Recombinant ADAMTS-13-loaded platelets localize to the site of thrombus formation in a more concentrated manner than enzyme replacement or plasma therapy. ADAMTS-13-encoding messenger RNA aims to induce a steady supply of secreted protein and gene therapy is a potentially curative strategy. Overall, targeted ADAMTS-13 replacement therapies may provide better outcomes than plasma therapy by achieving higher levels of ADAMTS-13 activity and a more sustained response with fewer adverse events. Herein, we describe targeted ADAMTS-13 replacement therapies for the treatment of TTP and discuss the advantages and limitations of each approach., Competing Interests: Declaration of competing interests None of the authors have any competing interests to disclose., (Copyright © 2023 International Society on Thrombosis and Haemostasis. Published by Elsevier Inc. All rights reserved.)

Parkinson's disease (PD) starts at the molecular and cellular level long before motor symptoms appear, yet there are no early-stage molecular biomarkers for diagnosis, prognosis prediction, or monitoring therapeutic response. This lack of biomarkers greatly impedes patient care and translational research-L-DOPA remains the standard of care more than 50 years after its introduction. Here, we performed a large-scale, multi-tissue, and multi-platform proteomics study to identify new biomarkers for early diagnosis and disease monitoring in PD. We analyzed 4877 cerebrospinal fluid, blood plasma, and urine samples from participants across seven cohorts using three orthogonal proteomics methods: Olink proximity extension assay, SomaScan aptamer precipitation assay, and liquid chromatography-mass spectrometry proteomics. We discovered that hundreds of proteins were upregulated in the CSF, blood, or urine of PD patients, prodromal PD patients with DAT deficit and REM sleep behavior disorder or anosmia, and non-manifesting genetic carriers of LRRK2 and GBA mutations. We nominate multiple novel hits across our analyses as promising markers of early PD, including DOPA decarboxylase (DDC), also known as L-aromatic acid decarboxylase (AADC), sulfatase-modifying factor 1 (SUMF1), dipeptidyl peptidase 2/7 (DPP7), glutamyl aminopeptidase (ENPEP), WAP four-disulfide core domain 2 (WFDC2), and others. DDC, which catalyzes the final step in dopamine synthesis, particularly stands out as a novel hit with a compelling mechanistic link to PD pathogenesis. DDC is consistently upregulated in the CSF and urine of treatment-naïve PD, prodromal PD, and GBA or LRRK2 carrier participants by all three proteomics methods. We show that CSF DDC levels correlate with clinical symptom severity in treatment-naïve PD patients and can be used to accurately diagnose PD and prodromal PD. This suggests that urine and CSF DDC could be a promising diagnostic and prognostic marker with utility in both clinical care and translational research., (© 2024. The Author(s).)

Background: Human prion diseases (HPDs) are fatal neurodegenerative disorders characterized by abnormal prion proteins (PrPSc). However, the detection of prion seeding activity in patients with high sensitivity remains challenging. Even though real-time quaking-induced conversion (RT-QuIC) assay is suitable for detecting prion seeding activity in a variety of specimens, it shows lower accuracy when whole blood, blood plasma, and blood-contaminated tissue samples are used. In this study, we developed a novel technology for the in vitro amplification of abnormal prion proteins in HPD to the end of enabling their detection with high sensitivity known as the enhanced quaking-induced conversion (eQuIC) assay., Methods: Three antibodies were used to develop the novel eQUIC method. Thereafter, SD50 seed activity was analyzed using brain tissue samples from patients with prion disease using the conventional RT-QUIC assay and the novel eQUIC assay. In addition, blood samples from six patients with solitary prion disease were analyzed using the novel eQuIC assay., Results: The eQuIC assay, involving the use of three types of human monoclonal antibodies, showed approximately 1000-fold higher sensitivity than the original RT-QuIC assay. However, when this assay was used to analyze blood samples from six patients with sporadic human prion disease, no prion activity was detected., Conclusion: The detection of prion seeding activity in blood samples from patients with sporadic prion disease remains challenging. Thus, the development of alternative methods other than RT-QuIC and eQuIC will be necessary for future research., (© 2024. The Author(s).)

Background: Exercise training promotes brain plasticity and is associated with protection against cognitive impairment and Alzheimer's disease (AD). These beneficial effects may be partly mediated by blood-borne factors. Here we used an in vitro model of AD to investigate effects of blood plasma from exercise-trained donors on neuronal viability, and an in vivo rat model of AD to test whether such plasma impacts cognitive function, amyloid pathology, and neurogenesis., Methods: Mouse hippocampal neuronal cells were exposed to AD-like stress using amyloid-β and treated with plasma collected from human male donors 3 h after a single bout of high-intensity exercise. For in vivo studies, blood was collected from exercise-trained young male Wistar rats (high-intensity intervals 5 days/week for 6 weeks). Transgenic AD rats (McGill-R-Thy1-APP) were injected 5 times/fortnight for 6 weeks at 2 months or 5 months of age with either (a) plasma from the exercise-trained rats, (b) plasma from sedentary rats, or (c) saline. Cognitive function, amyloid plaque pathology, and neurogenesis were assessed. The plasma used for the treatment was analyzed for 23 cytokines., Results: Plasma from exercised donors enhanced cell viability by 44.1% (p = 0.032) and reduced atrophy by 50.0% (p < 0.001) in amyloid-β-treated cells. In vivo exercised plasma treatment did not alter cognitive function or amyloid plaque pathology but did increase hippocampal neurogenesis by ∼3 fold, regardless of pathological stage, when compared to saline-treated rats. Concentrations of 7 cytokines were significantly reduced in exercised plasma compared to sedentary plasma., Conclusion: Our proof-of-concept study demonstrates that plasma from exercise-trained donors can protect neuronal cells in culture and promote adult hippocampal neurogenesis in the AD rat brain. This effect may be partly due to reduced pro-inflammatory signaling molecules in exercised plasma., (Copyright © 2023. Production and hosting by Elsevier B.V.)

Background: Applications of nonthermal plasma have expanded beyond the biomedical field to include antibacterial, anti-inflammatory, wound healing, and tissue regeneration. Plasma enhances epithelial cell repair; however, the potential damage to deep tissues and vascular structures remains under investigation., Result: This study assessed whether liquid plasma (LP) increased nitric oxide (NO) production in human umbilical vein endothelial cells by modulating endothelial NO synthase (eNOS) phosphorylation and potential signaling pathways. First, we developed a liquid plasma product and confirmed the angiogenic effect of LP using the Matrigel plug assay. We found that the NO content increased in plasma-treated water. NO in plasma-treated water promoted cell migration and angiogenesis in scratch and tube formation assays via vascular endothelial growth factor mRNA expression. In addition to endothelial cell proliferation and migration, LP influenced extracellular matrix metabolism and matrix metalloproteinase activity. These effects were abolished by treatment with NG-L-monomethyl arginine, a specific inhibitor of NO synthase. Furthermore, we investigated the signaling pathways mediating the phosphorylation and activation of eNOS in LP-treated cells and the role of LKB1-adenosine monophosphate-activated protein kinase in signaling. Downregulation of adenosine monophosphate-activated protein kinase by siRNA partially inhibited LP-induced eNOS phosphorylation, angiogenesis, and migration., Conclusion: The present study suggests that LP treatment may be a novel strategy for promoting angiogenesis in vascular damage. Video Abstract., (© 2024. The Author(s).)

Altered properties of fibrin clots have been associated with bleeding and thrombotic disorders, including hemophilia or trauma and heart attack or stroke. Clotting factors, such as thrombin and tissue factor, or blood plasma proteins, such as fibrinogen, play critical roles in fibrin network polymerization. The concentrations and combinations of these proteins affect the structure and stability of clots, which can lead to downstream complications. The present work includes clots made from plasma and purified fibrinogen and shows how varying fibrinogen and activation factor concentrations affect the fibrin properties under both conditions. We used a combination of scanning electron microscopy, confocal microscopy, and turbidimetry to analyze clot/fiber structure and polymerization. We quantified the structural and polymerization features and found similar trends with increasing/decreasing fibrinogen and thrombin concentrations for both purified fibrinogen and plasma clots. Using our compiled results, we were able to generate multiple linear regressions that predict structural and polymerization features using various fibrinogen and clotting agent concentrations. This study provides an analysis of structural and polymerization features of clots made with purified fibrinogen or plasma at various fibrinogen and clotting agent concentrations. Our results could be utilized to aid in interpreting results, designing future experiments, or developing relevant mathematical models.

Background: SLAMF1/CD150 is an active player in B cell signaling networks in chronic lymphocytic leukemia (CLL). CD150-mediated signaling initiates through a homophilic CD150 binding, which spans the adjacent cells, or the interaction with the soluble CD150 isoform (sCD150). The expression of sCD150 isoform at the mRNA and protein levels ex vivo was confirmed. However, it is unclear whether sCD150 isoform present in the blood plasma of CLL patients is a factor in the constitutive activation of CD150+ cells. The aim of this study was to develop an ELISA assay for the specific sCD150 evaluation and assess the sCD150 levels in the blood plasma of CLL patients with different CD150 expression on B cells., Materials and Methods: Blood plasma samples and peripheral blood mononuclear cells from 40 previously untreated CLL patients were analyzed. An ELISA method, ex vivo drug sensitivity assay, and a cell viability assay were used., Results: The sCD150 isoform was found in all studied plasma samples of CLL patients at different levels regardless of the cell surface CD150 expression status of B cells and sCD150 mRNA expression. CLL cases with low levels of the cell surface CD150 expression in B cells are characterized by high levels of sCD150 in blood plasma in contrast to the CLL cases with high cell surface CD150 expression on B cells. The elevated levels of sCD150 in blood plasma are associated with a better sensitivity of malignant B cells to cyclophosphamide and bendamustine., Conclusions: The sCD150 isoform is actively secreted by CLL B cells with its accumulation in blood plasma, which may be regarded as an additional factor in the CLL clinicopathologic variability.

Snakebite envenomation is classified as a Neglected Tropical Disease. Bothrops jararaca venom induces kidney injury and coagulopathy. HF3, a hemorrhagic metalloproteinase of B. jararaca venom, participates in the envenomation pathogenesis. We evaluated the effects of HF3 in mouse kidney and blood plasma after injection in the thigh muscle, mimicking a snakebite. Transcriptomic analysis showed differential expression of 31 and 137 genes related to kidney pathology after 2 h and 6 h, respectively. However, only subtle changes were observed in kidney proteome, with differential abundance of 15 proteins after 6 h, including kidney injury markers. N-terminomic analysis of kidney proteins showed 420 proteinase-generated peptides compatible with meprin specificity, indicating activation of host proteinases. Plasma analysis revealed differential abundance of 90 and 219 proteins, respectively, after 2 h and 6 h, including coagulation-cascade and complement-system components, and creatine-kinase, whereas a semi-specific search of N-terminal peptides indicated activation of endogenous proteinases. HF3 promoted host reactions, altering the gene expression and the proteolytic profile of kidney tissue, and inducing plasma proteome imbalance driven by changes in abundance and proteolysis. The overall response of the mouse underscores the systemic action of a hemorrhagic toxin that transcends local tissue damage and is related to known venom-induced systemic effects., Competing Interests: Declaration of competing interest The authors have no financial or non-financial interests to disclose. The authors declare that they have no conflicts of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The pathogenesis of Alzheimer's disease (AD) is believed to involve the accumulation of amyloid-β in the brain, which is produced by the sequential cleavage of amyloid precursor protein (APP) by β-secretase and γ-secretase. Recently, analysis of genomic DNA and mRNA from postmortem brain neurons has revealed intra-exonic recombinants of APP (gencDNA), which have been implicated in the accumulation of amyloid-β. In this study, we computationally analyzed publicly available sequence data (SRA) using probe sequences we constructed to screen APP gencDNAs. APP gencDNAs were detected in SRAs constructed from both genomic DNA and RNA obtained from the postmortem brain and in the SRA constructed from plasma cell-free mRNA (cf-mRNA). The SRA constructed from plasma cf-mRNA showed a significant difference in the number of APP gencDNA reads between SAD and NCI: the p-value from the Mann-Whitney U test was 5.14 × 10 -6 . The transcripts were also found in circulating nucleic acids (CNA) from our plasma samples with NGS analysis. These data indicate that transcripts of APP gencDNA can be detected in blood plasma and suggest the possibility of using them as blood biomarkers for Alzheimer's disease., (© 2023. The Author(s).)

Alzheimer's disease (AD) is the most common type of dementia, characterized by the abnormal accumulation of protein aggregates in the brain, known as neurofibrillary tangles and amyloid-β (Aβ) plaques. It is believed that an imbalance between cerebral and peripheral pools of Aβ may play a relevant role in the deposition of Aβ aggregates. Therefore, in this study, we aimed to evaluate the effect of the removal of Aβ from blood plasma on the accumulation of amyloid plaques in the brain. We performed monthly plasma exchange with a 5% mouse albumin solution in the APP/PS1 mouse model from 3 to 7 months old. At the endpoint, total Aβ levels were measured in the plasma, and soluble and insoluble brain fractions were analyzed using ELISA. Brains were also analyzed histologically for amyloid plaque burden, plaque size distributions, and gliosis. Our results showed a reduction in the levels of Aβ in the plasma and insoluble brain fractions. Interestingly, histological analysis showed a reduction in thioflavin-S (ThS) and amyloid immunoreactivity in the cortex and hippocampus, accompanied by a change in the size distribution of amyloid plaques, and a reduction in Iba1-positive cells. Our results provide preclinical evidence supporting the relevance of targeting Aβ in the periphery and reinforcing the potential use of plasma exchange as an alternative non-pharmacological strategy for slowing down AD pathogenesis.