The Arising Role of Extracellular Vesicles in Cholangiocarcinoma: A Rundown of the Current Knowledge Regarding Diagnostic and Therapeutic Approaches.

Cholangiocarcinomas (CCAs) constitute a heterogeneous group of highly malignant epithelial tumors arising from the biliary tree. This cluster of malignant tumors includes three distinct entities, the intrahepatic, perihilar, and distal CCAs, which are characterized by different epidemiological and molecular backgrounds, as well as prognosis and therapeutic approaches. The higher incidence of CCA over the last decades, the late diagnostic time that contributes to a high mortality and poor prognosis, as well as its chemoresistance, intensified the efforts of the scientific community for the development of novel diagnostic tools and therapeutic approaches. Extracellular vesicles (EVs) comprise highly heterogenic, multi-sized, membrane-enclosed nanostructures that are secreted by a large variety of cells via different routes of biogenesis. Their role in intercellular communication via their cargo that potentially contributes to disease development and progression, as well as their prospect as diagnostic biomarkers and therapeutic tools, has become the focus of interest of several current studies for several diseases, including CCA. The aim of this review is to give a rundown of the current knowledge regarding the emerging role of EVs in cholangiocarcinogenesis and their future perspectives as diagnostic and therapeutic tools.

Cardiometabolic factors and risk of non-arteritic anterior ischemic optic neuropathy: a systematic review and meta-analysis.

PURPOSE: The purpose of this systematic review and meta-analysis of the literature is to evaluate the association between cardiometabolic risk factors (hypertension, diabetes mellitus, hypercholesterolemia/dyslipidemia, HDL cholesterol, LDL cholesterol, lipoprotein(a), and triglycerides) and non-arteritic anterior ischemic optic neuropathy (NAION). METHODS: Pertinent publications were identified through a systematic search in PubMed and EMBASE databases, without language restrictions. The pooled odds ratios (OR) and standardized mean differences (SMD), with their 95% confidence intervals (95% CI) were estimated using random effects (DerSimonian Laird) models, as appropriate. A set of subgroup analyses and meta-regression analysis models were performed. RESULTS: Twenty-one studies (including 1560 patients with NAION and 2292 controls), examining the association between NAION and cardiometabolic risk factors, were eligible for the systematic review and meta-analysis. Hypertension (pooled OR = 1.50; 95% CI: 1.16-1.94), diabetes mellitus (pooled OR = 1.71; 95% CI: 1.33-2.21), and hypercholesterolemia/dyslipidemia (pooled OR = 2.00; 95% CI: 1.53-2.62) were associated with NAION. Among the components of dyslipidemia, higher serum triglycerides were associated with NAION, with a medium effect size (SMD = + 0.58, 95% CI: + 0.12 to + 1.04), whereas synthesis of four studies reporting on HDL and LDL cholesterol did not reveal any significant associations. A significant association between NAION and higher serum lipoprotein(a) levels (pooled OR = 2.88; 95%CI: 1.01-8.21) was also noted. CONCLUSIONS: This systematic review and meta-analysis found that NAION was associated with cardiometabolic factors, suggesting that vascular dysfunction may be implicated in the pathogenesis of the disease. Our findings may alert health care providers to try modifying these risk factors for NAION prevention.

The Emerging Role of Extracellular Vesicles and Autophagy Machinery in NASH-Future Horizons in NASH Management.

Non-alcoholic fatty liver disease (NAFLD) is considered the most frequent chronic hepatic disease in the general population, while it is the first cause of liver transplantation in the US. NAFLD patients will subsequently develop non-alcoholic steatohepatitis (NASH), which is characterized by aberrant hepatocellular inflammation with or without the presence of fibrosis. The lack of specific biomarkers and therapeutic strategies makes non-alcoholic steatohepatitis (NASH) management a difficult task for clinicians. Extracellular vesicles (EVs) constitute a heterogenic population of vesicles produced by inward or outward plasma-membrane budding. There is an emerging connection between autophagy EVs production, via an unconventional non-degradative procedure. Alterations in the amount of the secreted EVs and the cargo they carry are also involved in the disease progression and development of NASH. Autophagy constitutes a multistep lysosomal degradative pathway that reassures cell homeostasis and survival under stressful conditions, such as oxygen and energy deprivation. It prevents cellular damage by eliminating defected proteins or nοn-functional intracellular organelles. At the same time, it reassures the optimal conditions for the cells via a different mechanism that includes the removal of cargo via the secretion of EVs. Similarly, autophagy machinery is also associated with the pathogenetic mechanism of NAFLD, while it has a significant implication for the progression of the disease and the development of NASH. In this review, we will shed light on the interplay between autophagy and EVs in NASH, the emerging connection of EVs production with the autophagy pathway, and their possible manipulation for developing future therapeutic strategies for NASH.

Osmotic hemolysis is a donor-specific feature of red blood cells under various storage conditions and genetic backgrounds.

BACKGROUND: Transfusion research has recently focused on the discovery of red blood cell (RBC) storage capacity biomarkers and the elucidation of donor variation effects. This shift of focus can further strengthen personalization of transfusion therapy, by revealing probable links between donor biology, RBC storage lesion profile, and posttransfusion performance. STUDY DESIGN AND METHODS: We performed a paired correlation analysis of osmotic fragility in freshly drawn RBCs and during cold storage in different preservative solutions at weekly intervals until unit's expiration date (n = 231), or following 24 h reconstitution in allogeneic plasma (n = 32) from healthy controls or transfusion-dependent beta-thalassemia patients. RESULTS: We observed exceptional correlation profiles (r > 0.700, p < 10-5 in most cases) of RBC osmotic fragility in the ensemble of samples, as well as in subgroups characterized by distinct genetic backgrounds (sex, beta-thalassemia traits, glucose-6-phosphate dehydrogenase deficiency) and storage strategies (additive solutions, whole blood, RBC concentrates). The mean corpuscular fragility (MCF) of fresh and stored RBCs at each storage time significantly correlated with the MCF of stored RBCs measured at all subsequent time points of the storage period (e.g., MCF values of storage day 21 correlated with those of storage days 28, 35 and 42). A similar correlation profile was also observed between the osmotic hemolysis of fresh/stored RBCs before and following in vitro reconstitution in plasma from healthy controls or beta-thalassemia patients. CONCLUSION: Our findings highlighted the potential of osmotic fragility to serve as a donor-signature on RBCs at every step of any individual transfusion chain (donor, blood product, and probably, recipient).

Pathophysiological aspects of red blood cells in end-stage renal disease patients resistant to recombinant human erythropoietin therapy.

OBJECTIVE: Modified, bioreactive red blood cells (RBCs) and RBC-derived microvesicles (MVs) likely contribute to the hematological and cardiovascular complications in end-stage renal disease (ESRD). This study assesses the physiological profile of RBCs in patients with ESRD receiving standard or high doses of recombinant human erythropoietin (rhEPO). METHOD: Blood samples from twenty-eight patients under sustained hemodialysis, responsive, or not to standard rhEPO administration were examined for RBC morphology, fragility, hemolysis, redox status, removal signaling, membrane protein composition, and microvesiculation before and after dialysis. Acute effects of uremic plasma on RBC features were examined in vitro through reconstitution experiments. RESULTS: Overall, the ESRD RBCs were characterized by pathological levels of shape distortions, surface removal signaling, and membrane exovesiculation, but reduced fragility compared to healthy RBCs. Irreversible transformation of RBCs was found to be a function of baseline Hb concentration. The more toxic uremic context in non-responsive patients compared to rhEPO responders was blunted in part by the antioxidant, antihemolytic, and anti-apoptotic effects of high rhEPO doses, and probably, of serum uric acid. A selective lower expression of RBC membrane in complement regulators (CD59, clusterin) and of CD47 "marker-of-self" was detected in non-responders and responders, respectively. Evidence for different short-term dialysis effects and probably for a different erythrocyte vesiculation mechanism in rhEPO responsive compared to non-responsive patients was also revealed. CONCLUSION: Deregulation of RBC homeostasis might involve diverse molecular pathways driving erythrocyte signaling and removal in rhEPO non-responders compared to responsive patients.

Uric acid variation among regular blood donors is indicative of red blood cell susceptibility to storage lesion markers: A new hypothesis tested.

BACKGROUND: Oxidative stress orchestrates a significant part of the red blood cell (RBC) storage lesion. Considering the tremendous interdonor variability observed in the "storability," namely, the capacity of RBCs to sustain the storage lesion, this study aimed at the elucidation of donor-specific factors that affect the redox homeostasis during the storage of RBCs in standard systems. STUDY DESIGN AND METHODS: The hematologic profile of regular blood donors (n = 78) was evaluated by biochemical analysis of 48 different variables, including in vivo hemolysis and plasma oxidant and antioxidant factors and statistical analysis of the results. The possible effect of the uric acid (UA) variable on RBC storability was investigated in leukoreduced CPD/SAGM RBC units (n = 8) collected from donors exhibiting high or low prestorage levels of UA, throughout the storage period. RESULTS: Among the hematologic variables examined in vivo, cluster analysis grouped the donors according to their serum UA levels. Plasma antioxidant capacity, iron indexes, and protein carbonylation represented covariants of UA factor. RBCs prepared by low- or high-UA donors exhibited significant differences between them in spheroechinocytosis, supernatant antioxidant activity, and other RBC storage lesion-associated variables. CONCLUSION: UA exhibits a storability biomarker potential. Intrinsic variability in plasma UA levels might be related to the interdonor variability observed in the storage capacity of RBCs. A model for the antioxidant effect of UA during the RBC storage is currently proposed.

Serological Screening and Risk Factors Associated with Leishmania infantum Positivity in Newly Diagnosed HIV Patients in Greece.

A serological screening was conducted to detect IgG antibodies against Leishmania infantum (L. infantum) in newly diagnosed human immunodeficiency virus (HIV) patients in Greece. The study also examined potential risk factors and the agreement of commercially available serological methods. IgG antibodies against L. infantum were detected using enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescence antibody test (IFAT), and Western blot (WB). Out of 155 samples, 14 (9.0%) tested positive for IgG antibodies against L. infantum using at least two methods. Statistical analysis showed substantial agreement between WB and IFAT methods (Cohen's kappa = 0.75) but moderate overall agreement among the three methods (Fleiss' kappa = 0.42). Additionally, HIV+ intravenous drug users faced 3.55 times (p = 0.025) higher risk of testing positive for L. infantum IgG, positing that anthroponotic transmission between these patients is a plausible hypothesis based on existing literature. Non-invasive and cost-effective techniques are preferred to detect asymptomatic infections, and leishmaniasis screening should be conducted immediately after HIV diagnosis in endemic regions to enable prophylactic treatment for leishmaniasis in addition to antiretroviral therapy. To maximize sensitivity, performing at least two different serological methods for each patient is recommended.

Effect of silica-based mesoporous nanomaterials on human blood cells.

Different mesoporous nanomaterials (MSNs) are constantly being developed for a range of therapeutic purposes, but they invariably interact with blood components and may cause hazardous side effects. Therefore, when designing and developing nanoparticles for biomedical applications, hemocompatibility should be one of the primary goals to assess their toxicity at the cellular level of all blood components. The aim of this study was to evaluate the compatibility of human blood cells (erythrocytes, platelets, and leukocytes) after exposure to silica-based mesoporous nanomaterials that had been manufactured using the sol-gel method, with Ca and Ce as doping elements. The viability of lymphocytes and monocytes was unaffected by the presence of MSNs at any concentration. However, it was found that all nanomaterials, at all concentrations, reduced the viability of granulocytes. P-selectin expression of all MSNs at all concentrations was statistically significantly higher in platelet incubation on the first day of storage (day 1) compared to the control. When incubated with MSNs, preserved platelets displayed higher levels of iROS at all MSNs types and concentrations examined. Ce-containing MSNs presented a slightly better hemocompatibility, although it was also dose dependent. Further research is required to determine how the unique characteristics of MSNs may affect various blood components in order to design safe and effective MSNs for various biomedical applications.

Precision Oncology: Circulating Microvesicles as New Biomarkers in a Very Early Stage of Colorectal Cancer.

BACKGROUND: The release of microvesicles (MVs) is an essential phenomenon for inter-cellular signaling in health and disease. The role of MVs in cancer is multidimensional and includes cancer cell survival, proliferation, and invasion. In this prospective study, we analyzed MV levels in colorectal cancer patients and assessed the importance of MV release in early-stage colorectal cancer and survival. METHODS: This study included 98 patients and 15 controls. The characterization of MVs from human plasma was performed by flow cytometry using monoclonal antibodies. RESULTS: The levels of total MVs and MUC-1-positive, tissue factor (TF)-positive, and endothelial cell-derived MVs (EMVs) were statistically significantly higher in the colon cancer patients than in the controls (p < 0.001). Furthermore, the subgroup of patients with very early-stage colorectal cancer also had statistically significant differences in the levels of the abovementioned MVs compared to the controls (p < 0.01). Highly differentiated tumors had lower levels of MUC-1-positive MVs (p < 0.02), EMVs (p < 0.002), and EMV/TF combinations (p < 0.001) versus those with tumors with low/intermediate differentiation. CONCLUSIONS: Our data demonstrate that the analysis of circulating MV levels in plasma could possibly become a tool for the early diagnosis of colon cancer at a very early stage of the disease.

The time-course linkage between hemolysis, redox, and metabolic parameters during red blood cell storage with or without uric acid and ascorbic acid supplementation.

Oxidative phenomena are considered to lie at the root of the accelerated senescence observed in red blood cells (RBCs) stored under standard blood bank conditions. It was recently shown that the addition of uric (UA) and/or ascorbic acid (AA) to the preservative medium beneficially impacts the storability features of RBCs related to the handling of pro-oxidant triggers. This study constitutes the next step, aiming to examine the links between hemolysis, redox, and metabolic parameters in control and supplemented RBC units of different storage times. For this purpose, a paired correlation analysis of physiological and metabolism parameters was performed between early, middle, and late storage in each subgroup. Strong and repeated correlations were observed throughout storage in most hemolysis parameters, as well as in reactive oxygen species (ROS) and lipid peroxidation, suggesting that these features constitute donor-signatures, unaffected by the diverse storage solutions. Moreover, during storage, a general "dialogue" was observed between parameters of the same category (e.g., cell fragilities and hemolysis or lipid peroxidation and ROS), highlighting their interdependence. In all groups, extracellular antioxidant capacity, proteasomal activity, and glutathione precursors of preceding time points anticorrelated with oxidative stress lesions of upcoming ones. In the case of supplemented units, factors responsible for glutathione synthesis varied proportionally to the levels of glutathione itself. The current findings support that UA and AA addition reroutes the metabolism to induce glutathione production, and additionally provide mechanistic insight and footing to examine novel storage optimization strategies.

Stratification of ßSß+ Compound Heterozygotes Based on L-Glutamine Administration and RDW: Focusing on Disease Severity.

Sickle cell disease (SCD) is heterogeneous in terms of manifestation severity, even more so when in compound heterozygosity with beta-thalassemia. The aim of the present study was to stratify ßSß+ patient blood samples in a severity-dependent manner. Blood from thirty-two patients with HbS/ß-thalassemia compound heterozygosity was examined for several parameters (e.g., hemostasis, inflammation, redox equilibrium) against healthy controls. Additionally, SCD patients were a posteriori (a) categorized based on the L-glutamine dose and (b) clustered into high-/low-RDW subgroups. The patient cohort was characterized by anemia, inflammation, and elevated coagulation. Higher-dose administration of L-glutamine was associated with decreased markers of inflammation and oxidation (e.g., intracellular reactive oxygen species) and an altered coagulation profile. The higher-RDW group was characterized by increased hemolysis, elevated markers of inflammation and stress erythropoiesis, and oxidative phenomena (e.g., membrane-bound hemoglobin). Moreover, the levels of hemostasis parameters (e.g., D-Dimers) were greater compared to the lower-RDW subgroup. The administration of higher doses of L-glutamine along with hydroxyurea seems to attenuate several features in SCD patients, probably by enhancing antioxidant power. Moreover, anisocytosis may alter erythrocytes' coagulation processes and hemolytic propensity. This results in the disruption of the redox and pro-/anti-inflammatory equilibria, creating a positive feedback loop by inducing stress erythropoiesis and, thus, the occurrence of a mixed erythrocyte population.

Blood Cell-Derived Microvesicles in Hematological Diseases and beyond.

Microvesicles or ectosomes represent a major type of extracellular vesicles that are formed by outward budding of the plasma membrane. Typically, they are bigger than exosomes but smaller than apoptotic vesicles, although they may overlap with both in size and content. Their release by cells is a means to dispose redundant, damaged, or dangerous material; to repair membrane lesions; and, primarily, to mediate intercellular communication. By participating in these vital activities, microvesicles may impact a wide array of cell processes and, consequently, changes in their concentration or components have been associated with several pathologies. Of note, microvesicles released by leukocytes, red blood cells, and platelets, which constitute the vast majority of plasma microvesicles, change under a plethora of diseases affecting not only the hematological, but also the nervous, cardiovascular, and urinary systems, among others. In fact, there is evidence that microvesicles released by blood cells are significant contributors towards pathophysiological states, having inflammatory and/or coagulation and/or immunomodulatory arms, by either promoting or inhibiting the relative disease phenotypes. Consequently, even though microvesicles are typically considered to have adverse links with disease prognosis, progression, or outcomes, not infrequently, they exert protective roles in the affected cells. Based on these functional relations, microvesicles might represent promising disease biomarkers with diagnostic, monitoring, and therapeutic applications, equally to the more thoroughly studied exosomes. In the current review, we provide a summary of the features of microvesicles released by blood cells and their potential implication in hematological and non-hematological diseases.

Climate Changes Exacerbate the Spread of Ixodes ricinus and the Occurrence of Lyme Borreliosis and Tick-Borne Encephalitis in Europe-How Climate Models Are Used as a Risk Assessment Approach for Tick-Borne Diseases.

Climate change has influenced the transmission of a wide range of vector-borne diseases in Europe, which is a pressing public health challenge for the coming decades. Numerous theories have been developed in order to explain how tick-borne diseases are associated with climate change. These theories include higher proliferation rates, extended transmission season, changes in ecological balances, and climate-related migration of vectors, reservoir hosts, or human populations. Changes of the epidemiological pattern have potentially catastrophic consequences, resulting in increasing prevalence of tick-borne diseases. Thus, investigation of the relationship between climate change and tick-borne diseases is critical. In this regard, climate models that predict the ticks' geographical distribution changes can be used as a predicting tool. The aim of this review is to provide the current evidence regarding the contribution of the climatic changes to Lyme borreliosis (LB) disease and tick-borne encephalitis (TBE) and to present how computational models will advance our understanding of the relationship between climate change and tick-borne diseases in Europe.

Deciphering the Relationship Between Free and Vesicular Hemoglobin in Stored Red Blood Cell Units.

Red blood cells (RBCs) release hemoglobin (Hb)-containing extracellular vesicles (EVs) throughout their lifespan in the circulation, and especially during senescence, by spleen-facilitated vesiculation of their membrane. During ex vivo aging under blood bank conditions, the RBCs lose Hb, both in soluble form and inside EVs that accumulate as a part of storage lesion in the supernatant of the unit. Spontaneous hemolysis and vesiculation are increasingly promoted by the storage duration, but little is known about any physiological linkage between them. In the present study, we measured the levels of total extracellular and EV-enclosed Hb (EV-Hb) in units of whole blood (n = 36) or packed RBCs stored in either CPDA-1 (n = 99) or in CPD-SAGM additive solution (n = 46), in early, middle, and late storage. The spectrophotometry data were subjected to statistical analysis to detect possible correlation(s) between storage hemolysis and EV-Hb, as well as the threshold (if any) that determines the area of this dynamic association. It seems that the percentage of EV-Hb is negatively associated with hemolysis levels from middle storage onward by showing low to moderate correlation profiles in all strategies under investigation. Moreover, 0.17% storage hemolysis was determined as the potential cut-off, above which this inverse correlation is evident in non-leukoreduced CPDA units. Notably, RBC units with hemolysis levels > 0.17% are characterized by higher percentage of nanovesicles (<100 nm) over typical microvesicles (100-400 nm) compared with the lower hemolysis counterparts. Our results suggest an ordered loss of Hb during RBC accelerated aging that might fuel targeted research to elucidate its mechanistic basis.

Early and Late-Phase 24 h Responses of Stored Red Blood Cells to Recipient-Mimicking Conditions.

The 24-hour (24 h) post-transfusion survival of donor red blood cells (RBCs) is an important marker of transfusion efficacy. Nonetheless, within that period, donated RBCs may encounter challenges able to evoke rapid stress-responses. The aim of the present study was to assess the effect of exposure to plasma and body temperature upon stored RBCs under recipient-mimicking conditions in vitro from the first hours "post-transfusion" up to 24 h. For this purpose, packed RBCs from seven leukoreduced CPD/SAGM units were reconstituted with plasma of twenty-seven healthy individuals and incubated for 24 h at 37oC. Three units were additionally used to examine stress-responses in 3-hour intervals post mixing with plasma (n = 5) until 24 h. All experiments were performed in shortly-, medium-, and long-stored RBCs. Hemolysis, redox, morphology, membrane protein binding and vesiculation parameters were assessed. Even though spontaneous hemolysis was minimal post-reconstitution, it presented a time-dependent increase. A similar time-course profile was evident for the concentration of procoagulant extracellular vesicles and the osmotic fragility (shortly-stored RBCs). On the contrary, mechanical fragility and reactive oxygen species accumulation were characterized by increases in medium-stored RBCs, evident even from the first hours in the recipient-mimicking environment. Finally, exposure to plasma resulted in rapid improvement of morphology, especially in medium-stored RBCs. Overall, some RBC properties vary significantly during the first 24 h post-mixing, at levels different from both the storage ones and the standard end-of-24 h. Such findings may be useful for understanding the performance of RBCs and their possible clinical effects -especially on susceptible recipients- during the first hours post-transfusion.

Innate Variability in Physiological and Omics Aspects of the Beta Thalassemia Trait-Specific Donor Variation Effects.

The broad spectrum of beta-thalassemia (ßThal) mutations may result in mild reduction (ß ++), severe reduction (ß +) or complete absence (ß 0) of beta-globin synthesis. ßThal heterozygotes eligible for blood donation are "good storers" in terms of red blood cell (RBC) fragility, proteostasis and redox parameters of storage lesion. However, it has not been examined if heterogeneity in genetic backgrounds among ßThal-trait donors affects their RBC storability profile. For this purpose, a paired analysis of physiological and omics parameters was performed in freshly drawn blood and CPD/SAGM-stored RBCs donated by eligible volunteers of ß ++ (N = 4), ß + (N = 9) and ß 0 (N = 2) mutation-based phenotypes. Compared to ß +, ß ++ RBCs were characterized by significantly lower RDW and HbA2 but higher hematocrit, MCV and NADPH levels in vivo. Moreover, they had lower levels of reactive oxygen species and markers of oxidative stress, already from baseline. Interestingly, their lower myosin and arginase membrane levels were accompanied by increased cellular fragility and arginine values. Proteostasis markers (proteasomal activity and/or chaperoning-protein membrane-binding) seem to be also diminished in ß ++ as opposed to the other two phenotypic groups. Overall, despite the low number of samples in the sub-cohorts, it seems that the second level of genetic variability among the group of ßThal-trait donors is reflected not only in the physiological features of RBCs in vivo, but almost equally in their storability profiles. Mutations that only slightly affect the globin chain equilibrium direct RBCs towards phenotypes closer to the average control, at least in terms of fragility indices and proteostatic dynamics.

Supplementation with uric and ascorbic acid protects stored red blood cells through enhancement of non-enzymatic antioxidant activity and metabolic rewiring.

Redox imbalance and oxidative stress have emerged as generative causes of the structural and functional degradation of red blood cells (RBC) that happens during their hypothermic storage at blood banks. The aim of the present study was to examine whether the antioxidant enhancement of stored RBC units following uric (UA) and/or ascorbic acid (AA) supplementation can improve their storability as well as post-transfusion phenotypes and recovery by using in vitro and animal models, respectively. For this purpose, 34 leukoreduced CPD/SAGM RBC units were aseptically split in 4 satellite units each. UA, AA or their mixture were added in the three of them, while the fourth was used as control. Hemolysis as well as redox and metabolic parameters were studied in RBC units throughout storage. The addition of antioxidants maintained the quality parameters of stored RBCs, (e.g., hemolysis, calcium homeostasis) and furthermore, shielded them against oxidative defects by boosting extracellular and intracellular (e.g., reduced glutathione; GSH) antioxidant powers. Higher levels of GSH seemed to be obtained through distinct metabolic rewiring in the modified units: methionine-cysteine metabolism in UA samples and glutamine production in the other two groups. Oxidatively-induced hemolysis, reactive oxygen species accumulation and membrane lipid peroxidation were lower in all modifications compared to controls. Moreover, denatured/oxidized Hb binding to the membrane was minor, especially in the AA and mix treatments during middle storage. The treated RBC were able to cope against pro-oxidant triggers when found in a recipient mimicking environment in vitro, and retain control levels of 24h recovery in mice circulation. The currently presented study provides (a) a detailed picture of the effect of UA/AA administration upon stored RBCs and (b) insight into the differential metabolic rewiring when distinct antioxidant "enhancers" are used.

Red Blood Cell Abnormalities as the Mirror of SARS-CoV-2 Disease Severity: A Pilot Study.

PURPOSE: Unraveling the pathophysiology of COVID-19 disease is of crucial importance for designing treatment. The purpose of this study is to investigate the effects of the disease on erythrocytes (RBCs) and to correlate the findings with disease severity. MATERIALS AND METHODS: Hospitalized patients (n = 36) with COVID-19 and control group of healthy volunteers (n = 18) were included in the study. Demographic data, clinical, laboratory and chest Computed Tomography (CT) findings at time of admission were recorded. Laboratory measurements included: Hemoglobin (H b), indirect billirubin, LDH, D-Dimers, and plasma free hemoglobin (plasma free-Hb). On RBCs were performed: osmotic fragility (MCF), Free-Hb after mechanical stress (Free-Hb-MECH), intracellular RBC concentration of calcium ions (iCa2+), intracellular ROS (iROS), G6PD, intracellular active caspase-3 (RBC-caspase-3), IgG immunoglobulins (RBC-IgGs), which are bound on RBCs' senescent neo-antigen proteins and RBC surface phosphatidylserine (RBC-PS). RESULTS: The percentage of males was 50 and 66% and the mean age was 65.16 ± 14.24 and 66.33 ± 13.48 years among patients and controls respectively (mean ± SD, p = 0.78). Upon admission patients' PO2/FiO2 ratio was 305.92 ± 76.75 and distribution of infiltration extend on chest CT was: 0-25% (N = 19), 25-50%: (N = 7), and 50-75% (N = 9). Elevated hemolysis markers (LDH and plasma free-Hb) were observed in patients compared to the control group. Patients' RBCs were more sensitive to mechanical stress, and exhibited significantly elevated apoptotic markers (iCa2+, RBC-PS). Plasma free Hb levels correlated with the extend of pulmonary infiltrates on chest CT in COVID-19 patients. Surprisingly, patients' RBC-iROS were decreased, a finding possibly related with the increased G6PDH levels in this group, suggesting a possible compensatory mechanism against the virus. This compensatory mechanism seemed to be attenuated as pulmonary infiltrates on chest CT deteriorated. Furthermore, RBC-IgGs correlated with the severity of pulmonary CT imaging features as well as the abnormality of lung function, which are both associated with increased disease severity. Lastly, patients' D-Dimers correlated with RBC surface phosphatidylserine, implying a possible contribution of the red blood cells in the thrombotic diathesis associated with the SARS-CoV-2 disease. CONCLUSION: This pilot study suggests that SARS-CoV-2 infection has an effect on red blood cells and there seems to be an association between RBC markers and disease severity in these patients.

Coagulation Abnormalities in Renal Pathology of Chronic Kidney Disease: The Interplay between Blood Cells and Soluble Factors.

Coagulation abnormalities in renal pathology are associated with a high thrombotic and hemorrhagic risk. This study aims to investigate the hemostatic abnormalities that are related to the interaction between soluble coagulation factors and blood cells, and the effects of hemodialysis (HD) on it, in end stage renal disease (ESRD) patients. Thirty-two ESRD patients under HD treatment and fifteen healthy controls were included in the study. Whole blood samples from the healthy and ESRD subjects were collected before and after the HD session. Evaluation of coagulation included primary and secondary hemostasis screening tests, proteins of coagulation, fibrinolytic and inhibitory system, and ADAMTS-13 activity. Phosphatidylserine (PS) exposure and intracellular reactive oxygen species (iROS) levels were also examined in red blood cells and platelets, in addition to the platelet activation marker CD62P. Platelet function analysis showed pathological values in ESRD patients despite the increased levels of activation markers (PS, CD62P, iROS). Activities of most coagulation, fibrinolytic, and inhibitory system proteins were within the normal range, but HD triggered an increase in half of them. Additionally, the increased baseline levels of ADAMTS-13 inhibitor were further augmented by the dialysis session. Finally, pathological levels of PS and iROS were measured in red blood cells in close correlation with variations in several coagulation factors and platelet characteristics. This study provides evidence for a complex coagulation phenotype in ESRD. Signs of increased bleeding risk coexisted with prothrombotic features of soluble factors and blood cells in a general hyperfibrinolytic state. Hemodialysis seems to augment the prothrombotic potential, while the persisted platelet dysfunction might counteract the increased predisposition to thrombotic events post-dialysis. The interaction of red blood cells with platelets, the thrombus, the endothelium, the soluble components of the coagulation pathways, and the contribution of extracellular vesicles on hemostasis as well as the identification of the unknown origin ADAMTS-13 inhibitor deserve further investigation in uremia.

The Multi-Faced Extracellular Vesicles in the Plasma of Chronic Kidney Disease Patients.

Extracellular vesicles (EVs) are membrane-enclosed nanoparticles released by most cells in body fluids and extracellular matrix. They function as signal transducers in intercellular communication, contributing to the maintenance of cell and tissue integrity. EVs biogenesis is deregulated in various pathologies, in structural and functional connection to the pathophysiology of donor cells. Consequently, EVs are considered diagnostic and monitoring factors in many diseases. Despite consensus as to their activity in promoting coagulation and inflammation, there is evidence suggesting protective roles for EVs in stress states. Chronic kidney disease (CKD) patients are at high risk of developing cardiovascular defects. The pathophysiology, comorbidities, and treatment of CKD may individually and in synergy affect extracellular vesiculation in the kidney, endothelium, and blood cells. Oxidative and mechanical stresses, chronic inflammation, and deregulation of calcium and phosphate homeostasis are established stressors of EV release. EVs may affect the clinical severity of CKD by transferring biological response modifiers between renal, vascular, blood, and inflammatory cells. In this Review, we focus on EVs circulating in the plasma of CKD patients. We highlight some recent advances in the understanding of their biogenesis, the effects of dialysis, and pharmacological treatments on them and their potential impact on thrombosis and vascular defects. The strong interest of the scientific community to this exciting field of research may reveal hidden pieces in the pathophysiology of CKD and thus, innovative ways to treat it. Overcoming gaps in EV biology and technical difficulties related to their size and heterogeneity will define the success of the project.