Body Fluids Research Articles

Activity of exoglycosidases in blood, urine, cerebrospinal fluid, and vitreous humor in individuals who died from ethyl alcohol poisoning

Lysosomal exoglycosidases, namely α-mannosidase (MAN), α-fucosidase (FUC), β-galactosidase (GAL), and β-glucuronidase (GLU), are of crucial importance in breaking down the oligosaccharide chains of multiple glycoconjugates. Those enzymes liberate monosaccharides from non-reducing ends of oligosaccharide chains. In this study, we have aimed to assess the potential utility of MAN, FUC, GAL, and GLU activities as indicators of ethanol abuse in individuals who died from ethanol intoxication, while also investigating the mechanisms underlying their deaths. The study group comprised 22 fatal ethanol-intoxicated individuals, while the control group included 30 deceased individuals whose body fluids showed no traces of alcohol. We measured the activities (pKat/mL) of MAN, FUC, GAL, and GLU in the supernatants of blood, urine, cerebrospinal fluid as well as vitreous humor. The results indicated significantly lower activities of MAN (p = 0.003), FUC (p = 0.008), GAL (p = 0.014), and GLU (p = 0.004) in the urine of individuals poisoned by ethanol as compared to the control group. Additionally, there was a significantly lower activity of MAN in the vitreous of those affected by ethyl alcohol poisoning (p = 0.016).

Liquid Biopsy in Cancer Diagnosis-Treatment Methodology and Applications

The comprehensive review of liquid biopsy as a cutting-edge, non-invasive technique for cancer detection, diagnosis, and monitoring may be found in the document “Liquid Biopsy in Cancer Diagnosis – Treatment Methodology and Applications.” Via liquid biopsy, tumor-related indicators present in physiological fluids such as blood, urine, and saliva are examined, including exosomes, circulating tumor cells (CTCs), and circulating tumor DNA (ctDNA). Liquid biopsy enables regular, real-time sampling of tumor dynamics, in contrast to standard tissue biopsies, which are invasive and restricted to particular tumor regions. This renders it a potent instrument for the identification of cancer mutations and the tracking of tumor progression, particularly in cases of metastatic or elusive cancers. Technological developments like digital droplet PCR (ddPCR) and next-generation sequencing (NGS) have greatly improved the sensitivity and precision of ctDNA detection, making it possible to identify unusualexceptionally rare genetic mutations. This enhances the efficacy of tailored cancer treatment and monitoring. The genetic heterogeneity of cancers is captured by liquid biopsies, providing information on several tumor subclones that may be missed by traditional biopsies. The document also describes the use of different body fluids for different forms of cancer, such as head and neck and urological tumors, such as saliva, urine, and blood. Blood is the most often utilized medium for tracking genetic changes due to its ease of accessibility. Saliva and urine are useful in the detection of head and neck malignancies and bladder cancer, respectively. Liquid biopsy has several advantages, but it also has drawbacks, such as the requirement for standardized procedures, high prices, and differences in sensitivity and specificity between platforms.In order to overcome these obstacles and integrate liquid biopsy into standard clinical practice, the paper emphasizes the significance of more research and development. This will improve patient outcomes, early cancer detection, and personalized treatment.

Investigating the Predictive Biomethods. of Hypertension (HTN) in Human Foetal Astrocytes (HFAs) using Modified Proteomic Methods

Background: Hypertension (HTN) is a major risk factor for Cerebrovascular and cardiovas-cular diseases, causing premature deaths. Hence, there is an urgent need for predictive bi-omarkers to detect HTN before its onset. Methods • Cell Culture Technique: Growth of HFAs, followed by conversion of normal (A2) HFAs intoreactive (A1) type, by using ATP solution. • Confocal Microscopy: Quantification of Glial Fibrillary Acidic Protein (GFAP) in A1 and A2 HFAs, by using antibody ab 7260 with AF 488. Proteomics Methos • Extraction of proteins from A1 and A2 HFAs, using Single-pot, Sol-id-phase-enhanced Sample Preparation (SP3) protocol. • Liquid Chromatography- Mass Spectrometry techniques for obtaining the lev-els of calcium-gated proteins in A1 and A2 cell-solutions. Results • Micrographs obtained by Leica Microsystem indicated thicker filaments in A1 HFAs compared to A2 HFAs. • Confocal results showed higher levels of GFAP in A1 HFAs (P> 0.05). • Proteomics data analysed by PEAKS software revealed that proteins, such as GFAP were higher in reactive HFAs. Conclusions: The results showed higher levels of GFAP in A1 HFAs. Therefore, early detection of GFAP in human body fluids could be a reliable predictive biomarker for HTN

Fluorescence Label‐Free Doxorubicin Sensor Using Polystyrene Sulfonate as a Synthetic Receptor in Whispering Gallery Mode Microresonators

AbstractThe conventional quantification of Doxorubicin (DXR), a crucial anticancer drug, typically relies on complex and expensive techniques, limiting measurements beyond hospital settings and at different time intervals. Here, a label‐free fluorescence sensor is presented for DXR utilizing Whispering Gallery Mode (WGM) resonators. The resonators are based on fluorescent microparticles coated with a nanometer‐thick polystyrene sulfonate (PSS) layer serving as a synthetic receptor. The sensor demonstrates linear response for both position and width of the resonance peaks within the concentration range of 1–10 µg mL−1 of DXR of clinical interest. It also exhibits good selectivity against other chemotherapy drugs and can operate in complex biological fluids. The findings underscore the potential of WGM resonators as versatile multiparameter sensors for the sensitive and selective detection of small molecules, also in complex biological media.

Peristaltic Flow of Jeffrey Fluid in an Inclined Porous Tube with Multi-Thrombosis

The fact that multiple thrombosis opposes blood flow and flow is rare in blood vessels is refined to a normal condition with catheter application. The inner surface of the circular blood arteries may not be smooth in the majority of diseased instances. Additionally, the small blood vessel peristalsis mechanism is formed. In this paper, we discuss a mathematical model for the flow of biological fluid (blood) in a circular tube with multi-thrombosis considered under peristaltic wave propagation. The blood flow in this tube is restricted due to the many thromboses present, and the flow is redesigned with the aid of a catheter. We model this non-Newtonian blood flow issue for Jeffrey fluid. In most pathological cases, the inner surface of the circular blood vessels may not be smooth. Further, the mechanism of the peristalsis is established for small blood vessels. Because of this biological phenomenon, the peristaltic movement of Jeffrey fluid in a porous annulus with slip is examined. Equations that govern energy and momentum are solved exactly, and graphical interpretation is made using mathematical software. Streamline graphs show the many thromboses that increase in height. The wall shear stress graphs show a sinusoidally advancing wave with peaks and dips of different amplitudes. This tube's unique crest and trough amplitude are caused by the presence of multiple thromboses. We obtained the impact of the pressure gradient on the permeability parameter (β). The pressure gradient falls as the permeability parameter rises.

Circulating RKIP and pRKIP in Early-Stage Lung Cancer: Results from a Pilot Study

Background: Lung cancer (LC) is the leading cause of cancer-related deaths. Although low-dose computed tomography (LD-CT) reduces mortality, its clinical use is limited by cost, radiation, and false positives. Therefore, there is an urgent need for non-invasive and cost-effective biomarkers. The Raf Kinase Inhibitor Protein (RKIP) plays a crucial role in cancer development and progression and may also contribute to regulating the tumor–immune system axis. This protein has recently been described in biological fluids. Therefore, we conducted a pilot case–control study to assess RKIP and phosphorylated RKIP (pRKIP) levels in the urine and blood of LC patients. Methods: A novel enzyme linked immunosorbent assay (ELISA) assay was used to measure RKIP and pRKIP levels in urine and blood samples of two cohorts of LC patients and healthy controls (HSs). Furthermore, the biomarkers levels were correlated with tumor characteristics. Results: Serum, but not urine, levels of RKIP were significantly elevated in LC patients, distinguishing them from low- and high-risk healthy subjects with 93% and 74% accuracy, respectively. The RKIP/pRKIP ratio (RpR score) showed an accuracy of 90% and 79% in distinguishing LC patients from HS and HR-HS, respectively. Additionally, the RpR score correlated better with dimension, stage, and lymph node involvement in the tumor group. Conclusions: The serum RKIP and pRKIP profile may be a promising novel biomarker for early-stage LC.

Superior Electrochemical Sensor Application of Co3O4/C Heterostructure in Rapid Analysis of Anticancer Drug Palbociclib in Pharmaceutical Formulations and Biological Fluids.

In this work, we report a study examining how different salt concentrations affect the structure and electrochemical performance of two Co3O4/C materials designed for the fabrication of an easy, cheap, fast, safe, and useful electrochemical sensor for the detection of Palbociclib (PLB). Co3O4 nanoparticles were successfully created by encapsulating them in N-doped amorphous carbon matrices by using the molten salt-assisted approach. In this process, different amounts of potassium iodate and zeolitic imidazolate framework-12 (ZIF-12) were used, followed by pyrolysis at 800 °C. Optimum Co3O4 embedded porous carbon structures were obtained, and the composite with the highest electrochemical properties was modified to a glassy carbon electrode (GCE) surface for PLB detection. The linear response spanned from 1.0 to 5.0 μM, featuring a limit of detection (LOD) of 0.122 μM and a limit of quantification (LOQ) of 0.408 μM; the correlation coefficient was calculated as 0.995. The high sensitivity of the method in detecting PLB in pharmaceutical samples and human urine demonstrated its feasibility, with recovery percentages ranging from 99.3% to 101.3% and relative standard deviation (RSD) values of <3%. Therefore, this technique will make a significant contribution to monitoring and improving existing cancer treatment options.

Development of two ultra-sensitive UHPLC-QqQ-MS/MS methods for the simultaneous determination of hydroxyzine and its active metabolite (cetirizine) in human blood: applications to real cases of forensic toxicology.

Both postmortem toxicological and medical-forensic examinations are very important in the case of analyzing various types of chemical substances. Hydroxyzine (HZ) is a first-generation antihistamine drug with a sedative effect that disrupts cognitive function and affects the ability to drive motor vehicles. Enzymatic oxidation of the hydroxy-methyl group to the carboxyl group leads to the formation of its main metabolite-cetirizine (CZ). CZ is the active substance of antiallergic drugs. Because it does not cross the BBB (blood-brain barrier) easily, it is less likely to cause drowsiness or affect memory and impair cognitive function. Therefore, in criminal studies, it is often important what medication had been taken by a person involved, e.g., in a car accident, HZ or CZ. The analysis of both antihistamine drugs is challenging, as usually very low concentrations of the compound of interest need to be determined. Thus, an ultra-sensitive UHPLC-QqQ-MS/MS method was developed for simultaneous determination of HZ and CZ in biological fluid samples. The lower limit of quantification (LOQ) for HZ and CZ was calculated as 0.345 and 0.3696ng/mL, respectively. Together with a reduced sample volume to 200 μL, it makes the developed method suitable for a sensitive multidrug forensic toxicological analysis. Samples were extracted with simple and fast liquid-liquid extraction (ethyl acetate, pH 9). The present method for the determination of HZ and CZ in human blood proved to be simple, fast, selective, and sensitive. The quantification by LC-MS/MS was successfully applied to the samples coming from 28 authentic biological fluids (blood, urine, vitreous humor, bile and stomach content), both antemortem and postmortem. The performed studies confirm that the developed method is characterized by a high extraction efficiency. Its accuracy, reproducibility, simplicity, and selectivity suggest its application in clinical, toxicological, and forensic laboratories.

Multiparametric profiling of HER2-enriched extracellular vesicles in breast cancer using Single Extracellular VEsicle Nanoscopy

BackgroundPatients with HER2-positive breast cancer can significantly benefit from HER2-directed therapy – such as the monoclonal antibody trastuzumab. However, some patients can develop therapy resistance or change HER2 status. Thus, we urgently need new, noninvasive strategies to monitor patients frequently. Extracellular vesicles (EVs) secreted from tumor cells are emerging as potential biomarker candidates. These membrane-delimited nanoparticles harbor molecular signatures of their origin cells; report rapidly on changes to cellular status; and can be frequently sampled from accessible biofluids.ResultsUsing Single Extracellular VEsicle Nanoscopy (SEVEN) platform that combines affinity isolation of EVs with super-resolution microscopy, here we provide multiparametric characterization of EVs with ~ 8 nm precision and molecular sensitivity. We first interrogated cell culture EVs affinity-enriched in tetraspanins CD9, CD63, and CD81; these transmembrane proteins are commonly found on EV membranes. SEVEN robustly provided critical parameters of individual, tetraspanin-enriched EVs: concentration, size, shape, molecular cargo content, and heterogeneity. Trastuzumab-resistant cells (vs. trastuzumab-sensitive) secreted more EVs. Additionally, EVs from trastuzumab-resistant cells had lower tetraspanin density and higher HER2 density. We also evaluated EVs affinity-enriched in HER2; we found that these EVs (vs. tetraspanin-enriched) were larger and more elongated. We further optimized analytical sample processing to assess a rare population of HER2-enriched EVs from patient plasma. In breast cancer patients with elevated HER2 protein expression (vs. controls), HER2-enriched EVs had distinct characteristics including typically increased number of tetraspanin molecules and larger size. Importantly, these EVs were on average 25-fold more abundant compared to no cancer controls.ConclusionsSEVEN revealed unique characteristics of HER2-enriched EVs in cultured cells and complex biological fluid. In combination with current clinical approaches, this method is well poised to support precise therapeutic decisions.Graphical abstract

Development and validation of multiresidue analysis method for biomonitoring of pesticides and metabolites in human blood and urine by LC-QToF-MS

The widespread use of pesticides and their consequential presence in the environment is a growing concern due to the harmful health effects associated with pesticide exposure. For clinical and toxicology laboratories, a method for simultaneously determining these compounds and their metabolic products in body fluids, such as blood and urine, is important. In the present study, a rapid, sensitive and simultaneous LC-QToF-MS method for detecting multiclass pesticides and metabolites in blood and urine samples has been developed and validated. Four sample preparation procedures, protein precipitation and three different variants of QuEChERS-based extraction were evaluated to find a suitable, simple, and effective sample pretreatment technique. The final optimized sample preparation method (acetonitrile; 400 μl, MgSO4; 40 mg and NaCl; 10 mg) was validated for accuracy, precision, matrix effect, recovery, stability, carryover, and dilution integrity. Analyte recoveries ranged from 75.40 to 113.54 % while accuracy was evaluated in the range of 71.41–108.26 % and precision (%RSD) in the range of 0.01 %–16.85 %. The limit of quantification (LOQ) for all compounds was established in the range of 0.82–7.05 ng mL-1. The developed reliable, robust, and sensitive method was successfully applied for the quantification of target pesticides and metabolites in human blood and urine samples. Evaluated samples resulted in detection of eleven analytes (seven pesticides and four metabolites).

Role of Exosomes in Salivary Gland Tumors and Technological Advances in Their Assessment.

Backgroud: Salivary gland tumors (SGTs) are rare and diverse neoplasms, presenting significant challenges in diagnosis and management due to their rarity and complexity. Exosomes, lipid bilayer vesicles secreted by almost all cell types and present in all body fluids, have emerged as crucial intercellular communication agents. They play multifaceted roles in tumor biology, including modulating the tumor microenvironment, promoting metastasis, and influencing immune responses. Results: This review focuses on the role of exosomes in SGT, hypothesizing that novel diagnostic and therapeutic approaches can be developed by exploring the mechanisms through which exosomes influence tumor occurrence and progression. By understanding these mechanisms, we can leverage exosomes as diagnostic and prognostic biomarkers, and target them for therapeutic interventions. The exploration of exosome-mediated pathways contributing to tumor progression and metastasis could lead to more effective treatments, transforming the management of SGT and improving patient outcomes. Ongoing research aims to elucidate the specific cargo and signaling pathways involved in exosome-mediated tumorigenesis and to develop standardized techniques for exosome-based liquid biopsies in clinical settings. Conclusions: Exosome-based liquid biopsies have shown promise as non-invasive, real-time systemic profiling tools for tumor diagnostics and prognosis, offering significant potential for enhancing patient care through precision and personalized medicine. Methods like fluorescence, electrochemical, colorimetric, and surface plasmon resonance (SPR) biosensors, combined with artificial intelligence, improve exosome analysis, providing rapid, precise, and clinically valid cancer diagnostics for difficult-to-diagnose cancers.

A NOVEL ELECTROCHEMICAL BIOSENSOR BASED ON CREATININE DEIMINASE FOR CREATININE ANALYSIS

This work presents the development of a new electrochemical biosensor based on creatinine deiminase for the determination of creatinine concentration in biological samples. The optimal method for immobilizing bioselective elements on the electrode surface was selected, specifically the covalent binding of creatinine deiminase in a BSA matrix using glutaraldehyde. The biosensors prepared in this way exhibited the highest sensitivity to creatinine and high reproducibility in the fabrication of biomembranes. The biosensor demonstrated a low limit of detection of creatinine, high signal stability (RSD = 3%), significant storage stability, and a wide linear range, enabling the detection of both normal and pathological creatinine concentrations in biological fluids. The study also investigated the influence of working solution parameters, such as ionic strength, buffer capacity, pH, and protein content, on the biosensor’s performance. The main analytical characteristics of the proposed electrochemical biosensor indicate its potential use for monitoring creatinine concentration in biological samples.

Inside out: Exploring edible biocatalytic biosensors for health monitoring

Edible biosensors can measure a wide range of physiological and biochemical parameters, including temperature, pH, gases, gastrointestinal biomarkers, enzymes, hormones, glucose, and drug levels, providing real-time data. Edible biocatalytic biosensors represent a new frontier within healthcare technology available for remote medical diagnosis. The main challenges to develop edible biosensors are: i) finding edible materials (i.e. redox mediators, conductive materials, binders and biorecognition elements such as enzymes) complying with Food and Drug Administration (FDA), European Food Safety Authority (EFSA) and European Medicines Agency (EMEA) regulations; ii) developing bioelectronics able to operate in extreme working conditions such as low pH (∼pH 1.5 gastric fluids etc.), body temperature (between 37 °C and 40 °C) and highly viscous bodily fluids that may cause surface biofouling issues. Nowadays, advanced printing techniques can revolutionize the design and manufacturing of edible biocatalytic biosensors.This review outlines recent research on biomaterials suitable for creating edible biocatalytic biosensors, focusing on their electrochemical properties such as electrical conductivity and redox potential. It also examines biomaterials as substrates for printing and discusses various printing methods, highlighting challenges and perspectives for edible biocatalytic biosensors.

Metabolic landscaping of extracellular vesicles from body fluids by phosphatidylserine imprinted polymer enrichment and mass spectrometry analysis

Extracellular vesicles (EVs) are emerging as new source of biomarkers discovery in liquid biopsy due to their stabilization in body fluids, protected by phospholipid bilayers. However, the metabolomics study of EVs is very little reported due to the lack of efficient and high-throughput isolation methods for clinical samples. In this study, phosphatidylserine imprinted polymers were employed for rapid and efficient EVs isolation from five human body fluids, including plasma, urine, amniotic fluid, cerebrospinal fluid, and saliva. The isolated EVs were subsequently analyzed for metabolomic studies by high-resolution mass spectrometry. Metabolic landscaping was conducted between the body fluids and their EVs, indicating EVs contain a large number of metabolites that are completely specific to the body fluid source. Finally, quantitative metabolomic analysis of EVs was carried out with plasma samples of hepatocellular carcinoma. Several differentially expressed exosomal metabolites were revealed including the upregulation of sphingosine (d18:1), taurochenodeoxycholic acid (TCDCA), pipecolic acid (PA), and 4-hydroxynonenal (4-HNE) and down-regulation of piperine, caffeine, and indole. We believe the proposed methodology will provide a deeper understanding of the molecular composition and functions of EVs as an alternative source for biomarker discovery.

Exosome-based drug delivery systems for enhanced neurological therapeutics.

Exosomes are small extracellular vesicles naturally secreted by cells into body fluids, enriched with bioactive molecules such as RNAs, proteins, and lipids. These nanosized vesicles play a crucial role in physiological and pathological processes by facilitating intercellular communication and modulating cellular responses, particularly within the central nervous system (CNS). Their ability to cross the blood-brain barrier and reflect the characteristics of their parent cells makes exosomal cargo a promising candidate for biomarkers in the early diagnosis and clinical assessment of neurological conditions. This review offers a comprehensive overview of current knowledge on the characterization of mammalian-derived exosomes, their application as drug delivery systems for neurological disorders, and ongoing clinical trials involving exosome-loaded cargo. Despite their promising attributes, a significant challenge remains the lack of standardized isolation methods, as current techniques are often complex, costly, and require sophisticated equipment, affecting the scalability and affordability of exosome-based therapies. The review highlights the engineering potential of exosomes, emphasizing their ability to be customized for targeted therapeutic delivery through surface modification or conjugation. Future advancements in addressing these challenges and leveraging the unique properties of exosomes could lead to innovative and effective therapeutic strategies in neurology.

Soluble markers of viral rebound and post-treatment HIV control.

We focus on the different classes of biological molecules measurable in easily accessible bodily fluids that have the potential to serve as biomarkers for the HIV post-treatment controller (PTC) phenotype and/or the timing of viral rebound after stopping antiretroviral therapy (ART). Various viral components and host factors measurable in body fluids can play crucial roles in understanding and predicting the PTC phenotype. We review recent findings linking viral components, the quantitative and qualitative features of antibodies (including autologous HIV-specific antibodies), markers of inflammation and tissue damage, other host proteins (including hormones such as sex hormones), as well as metabolites, extracellular vesicles, and cell-free DNA to HIV control post-ART interruption. Several of these molecules can or have the potential to predict the time and probability of viral rebound after stopping ART and are biologically active molecules that can directly or indirectly (by modulating immune pressures) impact the size and activity of HIV reservoirs during and post-ART interruption. A comprehensive model combining multiple markers is needed to predict the PTC phenotype. This model can be leveraged to predict and understand the PTC phenotype, which can guide novel curative interventions to replicate this phenotype in post-treatment non-controllers.

Exploring the Experience of Guo Yi Master Lei Zhongyi in Dialectical Treatment of Heart Failure Based on the Theory of "Yang Wei Yin Xian"

The term "heart failure" first appeared in Wang Shuhe's "Meridian: Spleen and Stomach Diseases". The related description of heart failure is recorded in the "Huangdi Neijing", which is a disease mainly characterized by palpitations, asthma, and edema. It is often the terminal stage of chronic heart disease that cannot be cured. Its basic pathogenesis is prolonged illness with deficiency of heart qi, loss of nourishment in the heart, weakness in blood circulation, prolonged stagnation of the heart meridians, cessation of water consumption, and the generation of phlegm and dampness, resulting in heart failure. The theory of "Yang Wei Yin Xian" is a highly summarized understanding by Zhang Zhongjing of the etiology and pathogenesis of chest pain and heartache. Chinese medical master Lei Zhongyi has been practicing medicine for more than 60 years and has unique insights in the prevention and treatment of heart failure, with considerable clinical efficacy. He believes that heart failure belongs to the principle of deficiency and excess, and the theory of "yang micro yin string" is also applicable to this disease. Its onset is closely related to the pathological changes of substances such as qi, blood, and body fluids. The method of treating phlegm and blood stasis together should run through the treatment process, especially for patients who consume qi and yin in the later stage. Emphasis should be placed on promoting blood circulation and nourishing qi and yin, and the treatment should focus on both attacking and tonifying, with Qi and blood regulating simultaneously. Based on clinical experience, the original Yangxin Huoxue Tang has achieved significant clinical efficacy.

NPCoronaPredict: A Computational Pipeline for the Prediction of the Nanoparticle-Biomolecule Corona.

The corona of a nanoparticle immersed in a biological fluid is of key importance to its eventual fate and bioactivity in the environment or inside live tissues. It is critical to have insight into both the underlying bionano interactions and the corona composition to ensure biocompatibility of novel engineered nanomaterials. A prediction of these properties in silico requires the successful spanning of multiple orders of magnitude of both time and physical dimensions to produce results in a reasonable amount of time, necessitating the development of a multiscale modeling approach. Here, we present the NPCoronaPredict open-source software package: a suite of software tools to enable this prediction for complex multicomponent nanomaterials in essentially arbitrary biological fluids, or more generally any medium containing organic molecules. The package integrates several recent physics-based computational models and a library of both physics-based and data-driven parametrizations for nanomaterials and organic molecules. We describe the underlying theoretical background and the package functionality from the design of multicomponent NPs through to the evaluation of the corona.

Postmortem study of adrenomedullin and cortisol in femoral serum and pericardial fluid related to acute pulmonary edema.

Currently, various tools aid in determining the cause of death and the circumstances surrounding it. Thanatochemistry is one such method that provides insights into the physiopathological mechanisms of death and the behavior of specific biomarkers in different body fluids postmortem. Certain biomarkers, characterized by their stability and specificity to vital tissues like the lungs, are associated with mechanisms contributing to death, such as acute pulmonary edema (APE). This study aims to analyze the behavior of midregional pro-adrenomedullin (MR-proADM) and cortisol levels, measured in pericardial fluid and femoral serum, in relation to the severity of APE, categorized according to specific criteria. Samples were collected from a total of 92 corpses (77 males, 15 females) with a mean age of 56.7 ± 15.2 years. The severity of APE associated with the deaths was classified into three groups: slight or absent (n = 7; 8.6%), medium or moderate (n = 16; 19.8%), and intense (n = 58;71.6%).The determination of MR-proADM and cortisol levels was conducted using ELISA kits and an Immunoassay Analyzer, respectively. Our results reveal a significant increase in MR-proADM concentration with the severity of APE. Furthermore, a correlation was established between cortisol and MR-proADM concentrations in both pericardial fluid and femoral serum samples. This indicates that the severity of APE influences the production of ADM, regardless of the specific underlying pathophysiological mechanisms. Cortisol values were also found to be higher in the intense APE group compared to the moderate group.This study contributes to our understanding of the relationship between MR-proADM and cortisol, and the severity of APE, shedding light on potential applications in postmortem investigations.

Nanostructured Affinity Membrane to Isolate Extracellular Vesicles from Body Fluids for Diagnostics and Regenerative Medicine.

Electrospun regenerated cellulose (RC) nanofiber membranes were prepared starting from cellulose acetate (CA) with different degrees of substitution. The process was optimized to obtain continuous and uniformly sized CA fibers. After electrospinning, the CA membranes were heat-treated to increase their tensile strength before deacetylation to obtain regenerated cellulose (RC). Affinity membranes were obtained by functionalization, exploiting the hydroxyl groups on the cellulose backbone. 1,4-Butanediol-diglycidyl ether was used to introduce epoxy groups onto the membrane, which was further bioconjugated with the anti-CD63 antibody targeting the tetraspanin CD63 on the extracellular vesicle membrane surface. The highest ligand density was obtained with an anti-CD63 antibody concentration of 6.4 µg/mL when bioconjugation was performed in carbonate buffer. The resulting affinity membrane was tested for the adsorption of extracellular vesicles (EVs) from human platelet lysate, yielding a very promising binding capacity above 10 mg/mL and demonstrating the suitability of this approach.