Human Plasma Research Articles

ОЦЕНКА ЭФФЕКТИВНОСТИ ВАКЦИН И ДЕЗИНФЕКТАНТОВ ПРОТИВ КЛОСТРИДИОЗА КОРОВ НА МОЛОЧНЫХ КОМПЛЕКСАХ РЕСПУБЛИКИ БАШКОРТОСТАН

The purpose of the study is to evaluate the preventive effectiveness of the use of modern veterinary vaccines (One Shot Ultra 8, TOXIPRA Plus, Koglavax and Pastanarm 8) and the disinfectant Forbicida in comparison with hydrogen peroxide against clostridiosis of dairy cows, carried out in conditions of large dairy complexes of the Republic of Bashkortostan. Analyzes of biomaterial (internal organs and blood serum) of cows were carried out at the testing center of the State Budgetary Institution “Bashkir Research and Production Veterinary Laboratory” in compliance with regulated methods and in accordance with approved regulatory documents – international standards (GOST) and technical specifications (TU). Based on the laboratory tests carried out, we can conclude that without objective identification of the species of anaerobic bacteria clostridia, uncontrolled vaccination of livestock does not provide the necessary level of protection of the body of dairy cattle from this disease. According to the obtained microscopic studies of pathological materials, the most common cultures of clostridia cells are the species of perfringens (C. perfringens) and chauvoei (C. chauvoei), the latter of which is the causative agent of livestock emkara. In this regard, to maintain the required level of specific immunity protection of cows, monthly monitoring of antibodies to toxins remains, the level of which should be at least 40 % inhibition. However, the high effectiveness of specific prevention of clostridia is not achieved by any of the listed vaccines, which requires mandatory monitoring of the level of inhibition of antibodies in blood serum to identified types of clostridia and the introduction of strict sanitary treatment of livestock areas with disinfectant solutions. The use of a 10 % solution of Forbicida or 6 % hydrogen peroxide with an exposure of 24 and 3 hours, respectively, ensures 100 % death of pathogenic bacteria of the 3rd–4th group.

Influence of the polysaccharide capsule on virulence and fitness of Klebsiella pneumoniae

IntroductionThe capsular polysaccharide (CPS) of pathogenic bacteria is a critical virulence factor, often evading phagocytosis by host immune cells, while also interfering with the contact of the pathogen with host cells and contributing to biofilm formation. Klebsiella pneumoniae, a Gram-negative human pathogen associated with high antimicrobial resistances, produces 77 CPS serotypes. The CPS masks proteinaceous factors but also protects K. pneumoniae from uptake by host phagocytic cells and activation of the complement system. In addition to nosocomial, urinary tract and bloodstream infections or pneumonia hypervirulent strains have a highly mucoid phenotype and can cause soft tissue infections, liver abscesses, and meningitis as well. The CPS is therefore crucial for both escaping detection by the immune system and enhancing the virulence potential.MethodsIn this study, we generated a non-encapsulated mutant (Kpn2146∆wza) to observe how the CPS interferes with K. pneumoniae adhesion, survival in blood, and invasiveness in an experimental infection model.ResultsInfection of A549 lung epithelial cells showed similar adherence levels for the wild-type and non-capsulated strain, while our data showed a moderately higher internalization of Kpn2146Δwza when compared to the wild-type. In whole blood killing assays, we demonstrate that the K. pneumoniae capsule is essential for survival in human blood, protecting K. pneumoniae against recognition and clearance by the human immune system, as well as complement-mediated opsonization and killing. The non-encapsulated mutant, in contrast, was unable to survive in either whole blood or human plasma. Infections of Galleria mellonella larvae showed a significantly decreased virulence potential of the CPS-deficient mutant.DiscussionIn conclusion, our data indicate a crucial role of CPS in vivo.

Effect of Riociguat on Adenine-Induced Chronic Kidney Disease in Rats

Riociguat is a soluble guanylate cyclase (sGC) activator that increases the levels of cyclic guanosine monophosphate (cGMP). cGMP is known to play a key role in regulating kidney function. This research sought to investigate the possible protective effects of riociguat on the kidneys in the context of chronic kidney disease (CKD). CKD was induced in male Wistar rats through adenine administration. A total of 24 rats were allocated into four groups and administered treatments over a period of 35 days. Group 1 received a normal diet and a vehicle (carboxymethylcellulose (0.5%)), serving as the control. Group 2 received adenine (0.25% w/w) in the feed and a vehicle. Groups 3 and 4 received adenine in the feed (0.25% w/w) plus riociguat (3 mg/kg/day) and riociguat (10 mg/kg/day), respectively. Adenine administration significantly elevated systolic blood pressure, plasma creatinine, urea, and neutrophil gelatinase-associated lipocalin (NGAL). Furthermore, adenine reduced creatinine clearance and increased the urinary albumin-to-creatinine ratio and urinary N-Acetyl-β-D-Glucosaminidase (NAG). Histopathologically, adenine caused renal tubular necrosis and fibrosis. Furthermore, adenine elevated the plasma concentration of interleukins (IL-1β and IL-6) and tumor necrosis factor-alpha (TNF-α). Adenine significantly increased renal malondialdehyde (MDA) and reduced glutathione reductase (GR), superoxide dismutase (SOD), catalase (CAT), and total antioxidant capacity (TAC). Treatment with riociguat attenuated adenine-induced hypertension, improved kidney function, and ameliorated histopathological changes. Riociguat also reduced kidney injury markers, inflammation, and renal oxidative stress. The renoprotective effect of riociguat is probably due to anti-inflammatory and antioxidant actions. This indicates that riociguat may have the potential to slow the progression of kidney damage in chronic kidney disease (CKD).

БИОХИМИЧЕСКИЕ МАРКЕРЫ ДИСФУНКЦИИ ГЕПАТОБИЛИАРНОЙ СИСТЕМЫ У НОВОТЕЛЬНЫХ КОРОВ

The purpose of research is to study the biochemical parameters of the blood of cows to search for informative biomarkers for assessing hepatobiliary disorders in the early lactation period. Objectives: a comprehensive biochemical study of the blood of cows in the early lactation period and determination of the activity of liver and skeletal muscle enzymes as indicators characterizing liver cell damage. Experimental studies were carried out in 7 farms of the Altai Region. A total of 87 blood samples were collected from animals during the study period. The material for research was the blood serum of cows during the period up to 45 days of lactation. As a result of research, it was found that in cows with fatty liver degeneration there is a significant increase in the concentration of bilirubin in the blood - up to 13.72 µmol/l, a decrease in the concentration of albumin beyond the standard ranges to 26.4 g/l cholesterol to 1.74 mmol/l, compared to healthy animals. The level of triglycerides in the blood of sick cows decreases to 0.33 mmol/l, compared to healthy animals (0.47 mmol/l), however, these changes are within standard intervals (0.22–0.55 mmol/l). A study of circulating liver enzymes showed that in cows with fatty degeneration there is a significant increase in the enzymatic activity of aspartate aminotransferase (AST) to 148.3 U/l, glutamyltransferase (GGT) to 30.0 U/l and creatine kinase (CK) to 667.8 U /l, which is 1.95, 1.55 and 6.71 times more than in healthy animals, respectively. Thus, changes in the concentrations of bilirubin, cholesterol, albumin, AST and GGT can be used as a diagnostic algorithm that is highly informative when testing disorders of the functional state of the liver, which will allow for timely correction.

Intelligent Microfluidics for Plasma Separation: Integrating Computational Fluid Dynamics and Machine Learning for Optimized Microchannel Design

Efficient separation of blood plasma and Packed Cell Volume (PCV) is vital for rapid blood sensing and early disease detection, especially in point-of-care and resource-limited environments. Conventional centrifugation methods for separation are resource-intensive, time-consuming, and off-chip, necessitating innovative alternatives. This study introduces “Intelligent Microfluidics”, an ML-integrated microfluidic platform designed to optimize plasma separation through computational fluid dynamics (CFD) simulations. The trifurcation microchannel, modeled using COMSOL Multiphysics, achieved plasma yields of 90–95% across varying inflow velocities (0.0001–0.05 m/s). The input fluid parameters mimic the blood viscosity and density used with appropriate boundary conditions and fluid dynamics to optimize the designed microchannels. Eight supervised ML algorithms, including Artificial Neural Networks (ANN) and k-Nearest Neighbors (KNN), were employed to predict key performance parameters, with ANN achieving the highest predictive accuracy (R2 = 0.97). Unlike traditional methods, this platform demonstrates scalability, portability, and rapid diagnostic potential, revolutionizing clinical workflows by enabling efficient plasma separation for real-time, point-of-care diagnostics. By incorporating a detailed comparative analysis with previous studies, including computational efficiency, our work underscores the superior performance of ML-enhanced microfluidic systems. The platform’s robust and adaptable design is particularly promising for healthcare applications in remote or resource-constrained settings where rapid and reliable diagnostic tools are urgently needed. This novel approach establishes a foundation for developing next-generation, portable diagnostic technologies tailored to clinical demands.

Preparing a Liposome-Aided Drug Delivery System: The Entrapment and Release Profiles of Doxorubicin and 9-(N-Piperazinyl)-5-methyl-12(H)-quino [3,4-b][1,4]benzothiazinium Chloride with Human Serum Albumin

Background/Objectives: The principal aim of this work was to prepare a liposomal drug delivery system based on the commercial drug doxorubicin (DOX) and a budding agent with promising anticancer activity, 9-(N-piperazinyl)-5-methyl-12(H)-quino [3,4-b][1,4]benzothiazinium chloride (9-PBThACl). Methods: A spectrophotometric methodology was used to meticulously investigate the drug entrapment and release characteristics of the new liposomal complexes (L) based on dipalmitoylphosphatidylcholine (DPPC) with human serum albumin (HSA) and its defeated analog (dHSA). Results: The impact of the operational parameters (temperature and pH) on the liposome/drug(s)/(d)HSA, namely [LDPPC/9-PBThACl/DOX ]:(d)HSA] systems, as well as the polarity of the phospholipid bilayer, was examined. In order to compare the experimental findings, mathematical models were employed to specify the analytical factors controlling the process of drug release/potential drug release from liposomes. The observed variations in the drug encapsulation and release profiles were due to the combination of liposomal conjugates with human plasma protein. Conclusions: It was proven that changes in the environmental pH directly affect the percentage of drug entrapment in liposomes and the medicine release efficiency. Moreover, the grouping tendency of the liposomal combinations was investigated using a principal component analysis (PCA) and a hierarchical clustering analysis (HCA). Finally, an analysis of variance (ANOVA) confirmed the statistical impact of pH buffering and changing temperature factors on the drug release characteristics of liposomal conjugates.

Investigating the clinical significance of E2F5 expression in circulating extracellular vesicles in prostate carcinoma.

Prostate-specific antigen (PSA) based screening strategy has caused a marked improvement in prostate cancer detection and reduction in associated mortality. However, its specificity and sensitivity is not optimal for differentiating different forms of prostate cancer, resulting in overtreatment of indolent tumors. E2F5, a member of the family of transcription factors plays essential roles during many cellular processes. E2F5 amplification is a major event in PCa. A liquid biopsy is a minimally invasive procedure to investigate the cancer-related molecules in circulating tumor cells (CTCs), cell-free DNA, and extracellular vesicles (EVs). Serum and plasma are attractive sources of EV-based biomarkers as blood sample acquisition is a minimally invasive procedure. Given, the shortcomings of PSA as a serum marker, the suitability of E2F5 expression and chemical properties of EVs can be used for the diagnosis of clinically significant prostate cancer. To technically refine the current histopathology-based diagnosis of PCa by adding it to the molecular readouts specific to disease states and biochemical fingerprints of EVs. The study included 100 TRUS-biopsied tissues, 50 representing 4 Gleason grades and another 50 representing BPH. Expression of E2F5 was studied in all the qualified tissues by Immunohistochemistry (IHC). Blood from patients was collected and EVs were isolated and characterized. E2F5 was highly upregulated through all Gleason grades, the maximum being in Gleason 10(5 + 5) and the minimum in Gleason 7(3 + 4), as compared to BPH. The EVs isolated from blood plasma were within 30-200 nm in size. E2F5, being a transcription factor, is highly overexpressed in malignant biopsy tissues, through all Gleason grades. BPH tissues served as control samples. EVs from blood plasma might serve as a potential liquid biopsy marker for predicting disease progression and better prognosis.

Self-referenced Digital Spectral Chromatic Local Surface Plasmon Resonance in Ultrasensitive Severe Sepsis Interleukin-6 Detection.

Clinical monitoring of cytokines, such as interleukin-6 (IL-6), enables a timely diagnosis and can significantly improve patient prognosis. In this study, we developed a rapid, label-free, ultrasensitive, and low matrix-effect method called chromatic digital nanoplasmon-metry (cDiNM) to detect IL-6 in human blood plasma. Utilizing a multiple filter configuration, two nonadjacent specific transmission wavelength bands are extracted. One is centered within the full-width-at-half-maximum (fwhm) range where the local surface plasmon resonance (LSPR) response of the 80 nm gold nanoparticles (AuNPs) is strongest, while the other band is narrowed and blue-shifted from the peak to a region with minor intensity change. Scattering images of AuNPs passing through these two bands are then captured simultaneously and independently via the red and green channels of a color scientific complementary metal-oxide-semiconductor (sCMOS) camera. This configuration allows every AuNPs' spectral chromatic image contrast to be a self-referenced subtractive analysis LSPR and facilitates evaluation of their changes induced by the IL-6 binding across numerous individual AuNPs. This method achieves IL-6 detection in blood plasma within 45 min, requiring only 0.5 mL of a 10-fold diluted, label-free sample, with a limit of detection and quantification (LOD and LOQ) of less than 19.2 and 87.8 fg/mL, respectively, and a recovery rate of 96%. In summary, cDiNM provides rapid and accurate IL-6 monitoring with promising potential for clinical application in sepsis patient care.

Comparison of whole blood versus red blood cells and plasma to correct trauma-induced coagulopathy ex vivo.

Early resuscitation is based on platelet-poor components such as red blood cells and plasma (RBC + P), contributing to platelet dilution and worsening of trauma-induced coagulopathy (TIC). We aimed to compare the ability of cold-stored whole blood (WB) versus RBC + P as a single component to correct TIC. Blood samples were collected on admission from trauma patients who required activation of the major hemorrhage protocol at a single UK major trauma center in 2021/2022. Samples were spiked ex vivo with volumes equivalent to two, four, or eight units of WB or RBC + P stored for a maximum of 2 weeks. Thromboelastometry, platelet counting, and multiple electrode aggregometry (MEA) were performed. Samples from 20 adult trauma patients were analyzed. Median age was 32 years (27-42), 89% were male, 70% had platelet dysfunction (tissue factor-activated ROTEM [EXTEM]-tissue factor-activated ROTEM with cytochalasin D [FIBTEM] clot amplitude at 5 min [A5] ≤ 30 mm), 65% were coagulopathic (EXTEM A5 ≤ 40 mm), and 42% died. EXTEM-FIBTEM A5 was higher following spiking with WB than RBC + P (33 mm, 26-33, vs. 27 mm, 24-30, p < .001). WB-spiking corrected platelet dysfunction in 2 patient samples out of 20, whereas RBC + P increased the frequency of platelet dysfunction (1/20 sample) and TIC (4/20 samples). RBC + P was associated with a dose-dependent deterioration in rotational thromboelastometry (ROTEM) clot strength and dynamics, platelet count, and aggregation in response to multiple agonists compared with WB-spiking, which maintained or partially corrected these abnormalities. Compared with RBC + P, WB better preserves ex vivo platelet-related ROTEM parameters, platelet count, and aggregation, but does not fully correct these common derangements of TIC.

Investigation of the Effects of Selenium Against 4-Nonylphenol-induced Toxicity in Rat Testis.

The study aims to investigate whether selenium (Se) has a protective role against testicular toxicity induced by 4-nonylphenol (4-NP) in rats and reduces oxidative damage. For this purpose, 30 adult male Sprague-Dawley rats (250-300g/90days old) were divided into five equal groups: control, sham control, Se, 4-NP, and 4-NP + Se. The trial lasted 48days, with 4-NP administered at 125mg/kg/day and Se at 0.5mg/kg/day. The general microscopic examination of the testicular tissue involved measuring the diameters of seminiferous tubules, epithelial heights, and the density of stage XIV tubules in sections stained with the triple staining method. Caspase 3 and CX43 expressions were observed immunohistochemically, and the numbers of live-dead and normal-abnormal spermatozoa were recorded. The levels of malondialdehyde (MDA) and superoxide dismutase (SOD) were determined in blood serum and testicular tissue. At the end of the study, testicular toxicity due to 4-NP was demonstrated cytologically, histologically, histometrically, biochemically, and immunohistochemically. Se showed a positive effect against this toxicity, as evidenced by higher stage XIV tubule density in the 4-NP + Se group, lower caspase 3 levels compared to the 4-NP group, decreased MDA levels, increased SOD levels in serum and testicular tissue, and a higher count of live and normal spermatozoa. When used alone, Se may cause metabolic adverse effects, such as decreased live weight gain, reduced tubule diameter and epithelial height, and increased caspase 3 expression, depending on the dose and duration of use.

Detailed Profiling of Protein Corona Formed by Polydopamine Nanoparticles in Human Plasma.

The term protein corona (PC) indicates proteins adsorbed onto the surface of nanostructures exposed to biological media such as blood or serum. The analysis of the composition, evolution, and effect of the PC complexed with nanomaterials gained attention in recent years due to the importance of these parameters in determining the biological fate of nanostructures. In particular, the PC represents the first component of a nanomaterial interfacing with biological structures, dictating parameters such as nanoparticle internalization, immune response, bioavailability, and even toxicity. Polydopamine nanoparticles (PDNPs), obtained through the polymerization of dopamine, are "smart" materials characterized by high biocompatibility, high antioxidant capacities, high tunability and surface reactivity, biodegradability, and the ability to act as photothermal conversion agents when irradiated with a near-infrared (NIR) light source. Despite many interesting applications of PDNPs are currently described in the scientific literature, there is still no comprehensive analysis of the phenomenon of PC formation consequent to the exposure of these nanomaterials to biological media. Moreover, to date, the investigation of the effects of light irradiation of photothermally active nanomaterials on the composition and evolution of the associated PC has been extremely limited. With this work, we aim to provide for the first time an analysis of the phenomenon of PC formation associated with PDNPs, before and after NIR light stimulation. We characterized the PC formed following exposure to human plasma and analyzed the effects of several parameters on the overall PC composition and quantity, such as the PDNP size, presence of a surface functionalization, exposure time, and irradiation with an NIR laser, demonstrating that these parameters play a pivotal role in the resulting PC composition. Eventually, we showed that PDNPs exposed to human plasma have significantly different properties with respect to bare PDNPs, showing higher internalization rates in human glioblastoma cells, a higher light absorption value, and enhanced photothermal conversion abilities.

Quantification of mycophenolic acid in plasma by isotope dilution liquid chromatography-tandem mass spectrometry candidate reference method.

Accurate measurements of plasma mycophenolic acid (MPA) are essential for therapeutic drug monitoring in transplant recipients and autoimmune diseases. The performance of plasma mycophenolic acid routine methods remains highly variable that calls for a candidate reference measurement procedure (cRMP) to improve the standardization of plasma mycophenolic acid measurements. In this study, sample preparation was based on protein precipitation with methanol followed by further dilution. The mycophenolic acid was quantified by the isotope dilution liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS) with electrospray ionization in positive ion mode. According to the Clinical and Laboratory Standards Institute (CLSI) documents C62-A and C50-A, the basic analytical performance of the candidate reference method was verified, such as linearity, limit of quantification, matrix effect, precision, accuracy, and uncertainty. Moreover, the candidate reference measurement procedure was compared with the routine liquid chromatography-tandem mass spectrometry (LC-MS/MS) method in a clinical laboratory. Based on the data, the mycophenolic acid in human plasma was well detected by ID-LC-MS/MS. No apparent interferences were found with the mycophenolic acid measurement. The calibration curve for the mycophenolic acid was linear in the concentration range of 0.1-50μg/mL with a correlation coefficient of 0.9999 under the optimum experimental conditions. This method was sensitive because the low limit of quantitation (LOQ) was 0.05μg/mL. The recoveries of MPA were 98.11-98.95%. The intra-day and inter-day coefficients of variations (CV) of our method were ≤ 1.53% and ≤ 0.51%, respectively. No obvious matrix effect was observed. There was a good correlation between this method and the clinical routine LC-MS/MS method. To sum up, we established and validated a reliable plasma MPA method using ID-LC/MS/MS. The desirable accuracy and precision of this method enable it to serve as a promising cRMP to improve the standardization for plasma MPA routine measurements.

Evaluation of metabolism-associated proteins in abdominal aortic aneurysm.

Abdominal aortic aneurysm (AAA) development is inversely associated with diabetes mellitus. Targeting glucose metabolism seems to be a beneficial strategy for preventing AAA growth. Several metabolism-related factors are associated with AAA development. This study aims to analyse the expression of the so far unstudied proteins irisin, follistatin, activin A and ghrelin (ligand and receptor) in human and murine aneurysmal tissue, to assess the involvement of these pathways in AAA. Gene and protein expression was evaluated in aneurysmal and control tissue samples, by qPCR and immunohistochemistry. Vascular smooth muscle cells (VSMC) were studied in vitro for expression. Circulating levels of the proteins of interest in human plasma samples were evaluated by ELISA. In human aneurysmal tissue, the proteins of interest are were predominantly expressed by VSMCs in neovessels. Expression by human VSMCs was confirmed in vitro. In human plasma, the concentration of Irisin was higher in AAA (+/- diabetes) compared to controls. Patients with AAA and type 2 diabetes treated with metformin had lower levels of follistatin and ghrelin. This study demonstrates irisin, follistatin and ghrelin as interesting proteins to be studied in the context of the observed negative association between AAA development and type 2 diabetes.

Changes in Serum Inflammatory Markers and in Clinical Periodontal Condition After Non-Surgical Periodontal Treatment in Hypertensive Patients

Background: Chronic inflammatory disorders, such as periodontitis, may contribute to pro-hypertensive inflammation. Objectives: This study aimed to analyze changes in parameters for periodontitis, such as periodontal inflamed surface area (PISA) and serum inflammatory markers, following non-surgical periodontal treatment in hypertensive patients. Methods: A quasi-experimental pre-and-post study was conducted, involving 42 controlled hypertensive patients with periodontitis. The patients underwent periodontal assessment and tests, including complete blood count, glucose, triglycerides, HDL-C, LDL-C, and serum levels of inflammatory biomarkers. All patients received scaling and root planning treatment in a single session and were reevaluated one month after the treatment. Results: Post-treatment evaluations showed significant improvements in periodontal inflammation parameters, such as pocket depth, attachment level, bleeding on probing, and biofilm percentage, with statistically significant differences (p &lt; 0.001). There were decreases in serum VEGF levels (p &lt; 0.001) and reductions in PISA associated with declines in cytokine levels such as IL-10, IL-6, IL-12p70, IL-17A, and VEGF. PISA for IL-6 and IL-10 had a positive correlation before periodontal treatment and with IL-1β and IL-10 after treatment. Conclusions: Hypertensive patients with periodontitis who underwent non-surgical periodontal treatment showed improvements in their periodontal condition, a decrease in cytokine levels such as VEGF, and reductions in PISA associated with declines in cytokines such as IL-10, IL-6, IL-12p70, IL-17A, and VEGF. These findings confirm the role of inflammation in hypertensive patients with periodontitis.

Clinical repercussions after liberal volume replacement in the intraoperative period of cancer patients in a Hospital Unit in the Amazon

During the perioperative period, intravascular volume management becomes crucial to achieve better postoperative outcomes. Maintenance of euvolemia is critical to ensure adequate tissue perfusion, since hypervolemia has been associated with higher rates of postoperative morbidity. One of the main challenges in managing surgical volume lies in intravascular monitoring. Despite the use of essential tools such as non-invasive blood pressure cuffs and heart rate monitoring, significant challenges persist due to difficulties in accurately measuring fluid losses, varied cardiovascular responses to anesthesia, and the absence of standardized assessment protocols for anesthetized patients. Given the critical role of adequate volume replacement in cancer patients undergoing medium-sized urologic surgeries, this study aims to identify potential adverse clinical consequences associated with volume replacement therapy in postoperative ICU patients undergoing radical prostatectomy and radical nephrectomy in an oncology referral center in the Brazilian Amazon. This prospective, descriptive and exploratory study will be carried out from August 2023 to August 2024 at the Amazonas Oncological Control Center Foundation (FCECON). The research seeks to elucidate specific adverse clinical outcomes related to fluid management in cancer patients undergoing these surgical procedures.

Metabolic Characteristics of Lame Cows During Puerperium and the Beginning of the Reproductive Period

This study presents findings from two discrete experimental processes that examined the impact of lameness events on two consecutive, critical time points in the annual production cycle of dairy cattle (early in puerperium—first study, and later at the onset of the reproductive period—second study) regarding liver function, glucose levels, milk production, body condition score, and back fat thickness. In the first study, 47 cows (lame group n = 22, control group n = 25) were monitored from 10 days ante partum (ap) to 46 days post-partum (pp). In the second study, 79 cows (lame group n = 52, control group n = 27) were monitored from day 28 ± 5 pp to day 65–72 ± 5 pp. Lame cows had greater gamma-glutamyl transferase (GGT) concentrations in the blood serum compared to control cows (25.83 vs. 23.56, p = 0.02, respectively) early in puerperium, whereas the two groups did not differ in the second study. The concentration of glutamate dehydrogenase (GLDH) was lower for lame compared to control cows in both studies (17.24 vs. 24.60, respectively, p = 0.02 in the first study, and 30.50 vs. 51.10, respectively, p = 0.02 in the second study). The concentrations of aspartate transaminase (AST) and glucose did not differ between groups in both studies. Lame cows had a lower body condition score (BCS) and backfat thickness (BFT) scores compared to the control in both studies overall. The lame cows of the first study experienced a significant loss of milk production, especially during the second month of lactation, while in the second study, milk production remained unaffected. Conclusively, lame cows have lower BCS and BFT values, whereas milk yield can be negatively affected only if lameness occurs early in the puerperium, probably beginning at the dry period. However, the current research shows that acutely lame cows, as described in this study, exhibit only mild alterations in liver function compared to non-lame ones.

Ectopic expression of the cation-chloride cotransporter KCC2 in blood exosomes as a biomarker for functional rehabilitation

BackgroundTraumatic brain injury (TBI) is a major cause of disabilities in industrialized countries. Cognitive decline typically occurs in the chronic phase of the condition, following cellular and molecular processes. In this study, we described the use of KCC2, a neuronal-specific potassium–chloride cotransporter, as a potent biomarker to predict cognitive dysfunction after TBI.MethodsUsing neuronal and total exosome collections from the blood serum of the controls and patients with TBI, we were able to anticipate the decline in cognitive performance.ResultsAfter TBI, we observed a significant and persistent loss of KCC2 expression in the blood exosomes, which was correlated with the changes in the network activity and cellular processes such as secondary neurogenesis. Furthermore, we established a correlation between this decrease in KCC2 expression and the long-term consequences of brain trauma and identified a link between the loss of KCC2 expression and the emergence of depressive-like behavior observed in the mice.ConclusionWe successfully validated our previous findings, supporting the potential therapeutic benefits of bumetanide in mitigating post-traumatic depression (PTD) following TBI. This effect was correlated with the recovery of KCC2 expression in the blood exosomes, the prevention of extensive neuronal loss among the interneurons, and changes in secondary neurogenesis.

Development of an atherosclerosis rabbit model to evaluate the hemodynamic impact of extracorporeal circulation.

Aortic atherosclerosis increases the risk of embolic events under extracorporeal circulation (ECC). To evaluate the hemodynamic impact of ECC on atheromatous plaques, an atherosclerosis animal model, which is also eligible for ECC, is required. Twenty-nine New Zealand White rabbits received a pro-atherosclerotic diet (group diet, n = 10), a pro-atherosclerotic diet and additional intraaortic balloon insufflation injury (group BI, n = 9), or served as controls (n = 10). After 3 or 6 months, aortic explants were analyzed by (immuno-)histology and RT-PCR. Blood serum analyses revealed increased cholesterol-levels in groups diet and BI compared to controls (3 months: p = 0.03 each, 6 months: p < 0.0001 each). Aortic inflammatory infiltration was significantly enhanced in groups diet (CD3 at 3 months: p < 0.0001, 6 months: p = 0.02; CD68 at 3 months: p = 0.01) and BI (CD3 at 3 months: p < 0.0001, 6 months: p = 0.03; CD68 at 3 months: p = 0.04, 6 months: p = 0.02). Increased intima hyperplasia occurred in both groups (p < 0.0001 each). Macroscopic analyses after 3 and 6 months showed ubiquitous lumen-narrowing aortic plaques. Calcification of the intima and media was increased in groups diet (intima: p < 0.0001 at 3 and 6 months; media at 3 months: p < 0.0001, 6 months: p = 0.01) and BI (intima: p < 0.0001 at 3 and 6 months; media at 3 months: p < 0.0001, 6 months: p = 0.02). Extensive lipid accumulation was found in the intima in both treatment groups (p < 0.0001 each). A rabbit model with high aortic calcific plaque burden-diet-induced with no implicit need of an additional intimal injury by an intraaortic balloon insufflation due to comparable outcome-exhibiting multiple pathophysiological aspects of human atherosclerosis has been designed and thoroughly characterized. It is suitable for use in future studies on the interaction between atherosclerotic plaques and the arterial blood flow under ECC.

High-Efficiency Capture and Proteomic Analysis of Plasma-Derived Extracellular Vesicles through Affinity Purification.

Plasma-derived extracellular vesicles (EVs) are promising sources of biomarkers. It is still a challenge to isolate EVs from a small amount of human plasma for downstream proteomic analysis. The isolation process is hindered by contamination with high-abundance blood proteins and lipoprotein particles, which adversely impact proteomic analyses. Moreover, although EV immune-isolation via magnetic beads often integrates with flow sorting and Western blotting (WB), it lacks compatibility with nanoparticle tracking analysis (NTA) and proteomic analysis. To address these issues, we have developed a functional affinity magnetic bead, EVlent (Extracellular Vesicles isoLated Efficiently, Naturally, and Totally), enabling the rapid and efficient isolation of EVs from plasma. By optimizing the quantities of magnetic beads and plasma used, we characterized the isolated EVs through WB, NTA, and transmission electron microscopy (TEM), showing the successful isolation of EVs from plasma. Proteomic analysis of these EVs identified over 2000 proteins and 15,000 peptides from 100 μL of plasma and nearly 1000 proteins from trace samples as small as 5 μL. Additionally, this isolation method significantly reduced contaminants, including plasma proteins and lipoproteins, compared to ultracentrifugation. Finally, we applied this strategy to plasma samples of healthy individuals and those with Parkinson's disease, identifying four potential biomarkers that provide promising guidance for clinical diagnosis.

Purified Monascus Pigments: Biological Activities and Mechanisms of Action.

Monascus pigments having yellow, orange, and red colors are widely studied for their potential beneficial properties. Many different biological activities have been reported regarding Monascus pigments and their derivatives, but the usual method is to test complex extracts from the mycelium of the fungus or from a fungus-fermented substrate. However, this review is mainly concerned with the biological activities of purified Monascus pigments. Both yellow (ankaflavin, monascin) and red (rubropunctamine, monascorubramine) Monascus pigments are proven antioxidants if used in concentrations of 10 μg/mL or higher. Antimicrobial activity against Gram-positive and Gram-negative bacteria and fungi has been observed with all Monascus pigments. However, the best antimicrobials are red Monascus pigments, and their amino acid derivatives (l-cysteine derivatives have MIC 4 μg/mL against Enterococcus faecalis). Yellow monaphilones and orange monaphilols seem to have the highest anti-inflammatory activity (IC50 1.7 μM of monaphilol D) and, together with red Monascus pigment derivatives, have mild antiobesity and antidiabetic activities. Further, monascin and ankaflavin in daily doses of 0.5 and 0.08 mg, respectively, lowered serum blood levels of low-density lipoprotein cholesterol complexes in rats on a high-fat diet. Orange Monascus pigments, rubropunctatin and monaphilols A and C, exhibit cytotoxic and antitumor activities (IC50 8-10 μM).