Leucoreduction helps to preserve activity of antioxidant barrier enzymes in stored red blood cell concentrates.

BACKGROUND: Oxidoreductive imbalance is a major cause of excessive haemolysis in in vitro conditions. Leucocytes and blood platelets present in red blood cell concentrates (RBCs) are one of the sources of free radicals, which have a significant effect on the status of stored erythrocytes. The study objective was to assess the effect of leucoreduction on the intensity of lipid peroxidation and the activity of antioxidant barrier enzymes in RBC. STUDY DESIGN AND METHODS: Red blood cell concentrates units obtained from 10 whole-blood units were split into two equal units, one of which was leucoreduced on the day of donation. Both units were stored for 35 days. The following markers of oxidoreductive balance were measured on day 0 (donation day) and on storage days 7, 14, 21 and 35: concentration of malondialdehyde (MDA) and activities of antioxidant barrier components, that is superoxide dismutase, glutathione peroxidase and glutathione reductase. RESULTS: Lipid peroxidation in leucodepleted units (LRBC) was slower than that in non-leucodepleted ones. The analysis of LRBC revealed statistically significant decrease in concentrations of MDA. The activities of superoxide dismutase, glutathione peroxidase and glutathione reductase were higher throughout the storage period as compared to non-leucoreduced RBC. Statistically significant differences between RBC and LRBC units were noted throughout the storage in the activity of lactate dehydrogenase, and concentrations of K(+) ions and free haemoglobin. CONCLUSIONS: Leucoreduction of RBC before storage helps to preserve the activity of antioxidant barrier enzymes in stored RBCs and significantly improves the quality of stored red blood cell components.

Decreased plasma thiol antioxidant barrier and selenoproteins as potential biomarkers for ongoing methylmercury intoxication and an individual protective capacity.

Manifestation of methylmercury (MeHg) toxicity depends on individual susceptibility to MeHg, as well as MeHg burden level. Therefore, biomarkers that reflect the protective capacity against MeHg are needed. The critical role of oxidative stress in the pathogenesis of MeHg cytotoxicity has been demonstrated. Because MeHg has high affinity for selenohydryl groups, sulfhydryl groups, and selenides, and causes posttranscriptional defects in selenoenzymes, proteins with selenohydryl and sulfhydryl groups should play a critical role in mediating MeHg-induced oxidative stress. Here, plasma oxidative stress markers and selenoproteins were investigated in MeHg-intoxicated rats showing neuropathological changes after 4 weeks of MeHg exposure. The thiol antioxidant barrier (-SHp) level significantly decreased 2 weeks after MeHg exposure, which is an early stage at which no systemic oxidative stress, histopathological changes, or clinical signs were detected. Diacron reactive oxidant metabolite (d-ROM) levels significantly increased 3 weeks after MeHg exposure, indicating the occurrence of systemic oxidative stress. Rats treated with lead acetate or cadmium chloride showed no changes in levels of -SHp and d-ROM. Selenoprotein P1 abundance significantly decreased in MeHg-treated rats, whereas it significantly increased in rats treated with Pb or Cd. Plasma selenium-dependent glutathione peroxidase (GPx3) activity also significantly decreased after MeHg exposure, whereas plasma non-selenoenzyme glutathione reductase activity significantly increased in MeHg-treated rats. The results suggest that decreased capacity of -SHp and selenoproteins (GPx3 and selenoprotein P) can be useful biomarkers of ongoing MeHg cytotoxicity and the individual protective capacity against the MeHg body burden.

Simulated Microgravity Exposure Induces Antioxidant Barrier Deregulation and Mitochondria Enlargement in TCam-2 Cell Spheroids.

One of the hallmarks of microgravity-induced effects in several cellular models is represented by the alteration of oxidative balance with the consequent accumulation of reactive oxygen species (ROS). It is well known that male germ cells are sensitive to oxidative stress and to changes in gravitational force, even though published data on germ cell models are scarce. We previously studied the effects of simulated microgravity (s-microgravity) on a 2D cultured TCam-2 seminoma-derived cell line, considered the only human cell line available to study in vitro mitotically active human male germ cells. In this study, we used a corresponding TCam-2 3D cell culture model that mimics cell-cell contacts in organ tissue to test the possible effects induced by s-microgravity exposure. TCam-2 cell spheroids were cultured for 24 h under unitary gravity (Ctr) or s-microgravity conditions, the latter obtained using a random positioning machine (RPM). A significant increase in intracellular ROS and mitochondria superoxide anion levels was observed after RPM exposure. In line with these results, a trend of protein and lipid oxidation increase and increased pCAMKII expression levels were observed after RPM exposure. The ultrastructural analysis via transmission electron microscopy revealed that RPM-exposed mitochondria appeared enlarged and, even if seldom, disrupted. Notably, even the expression of the main enzymes involved in the redox homeostasis appears modulated by RPM exposure in a compensatory way, with GPX1, NCF1, and CYBB being downregulated, whereas NOX4 and HMOX1 are upregulated. Interestingly, HMOX1 is involved in the heme catabolism of mitochondria cytochromes, and therefore the positive modulation of this marker can be associated with the observed mitochondria alteration. Altogether, these data demonstrate TCam-2 spheroid sensitivity to acute s-microgravity exposure and indicate the capability of these cells to trigger compensatory mechanisms that allow them to overcome the exposure to altered gravitational force.

Oxidative Stress Evaluation in Dogs Affected with Canine Monocytic Ehrlichiosis.

The study aimed to evaluate the concentration of reactive oxidative metabolites (R-OOHs), the antioxidant barrier (OXY), and the ratio between R-OOHs and OXY (OSi) and thiol groups of plasma compounds (SHp) in in canine monocytic ehrlichiosis. Thirty dogs affected with monocytic ehrlichiosis (canine monocytic ehrlichiosis group-CME group) and ten healthy dogs (control group-CTR group) were evaluated. CME was diagnosed by the presence of clinical signs and the detection of anti-Ehlichia canis antibodies. Oxidative stress parameters of two groups were compared using the Mann-Whitney test. Significance was set at p < 0.05. Spearman rank correlation was performed to analyze oxidative stress, and hematological and biochemical variables in the CME group. All dogs affected with CME showed a wide spectrum of clinical signs such as lethargy, anorexia, fever, weight loss, lymph adenomegaly, splenomegaly, subcutaneous and mucosal petechial and ecchymosis, and vomiting. Anaemia, leukocytosis, thrombocytopenia, hyperglobulinemia, hypoalbuminemia and an increase of blood urea nitrogen and creatinine are also detected. Results showed significantly lower values of SHp in the CME group than in CTR. A statistically significant difference in the number of white blood cells, platelets, and blood urea nitrogen concentration was assayed comparing to the two groups. A negative correlation between SHp and hemoglobin concentration was recorded. These preliminary results may suggest a possible function of oxidative stress in the onset of clinical signs during the course of CME.

Oxidative Stress and High-Mobility Group Box 1 Assay in Dogs with Gastrointestinal Parasites.

This study aimed to evaluate the concentration of reactive oxidative metabolites, the antioxidant barrier, thiol groups of plasma compounds, and high-mobility group box 1 in shelter dogs naturally infected with helminths. In addition, the correlation between clinical signs and oxidative stress was investigated. Sixty-six (41 male and 25 female) adult mixed-breed dogs housed in a shelter with the diagnosis of gastrointestinal nematodes (i.e., Ancylostoma spp., Uncinaria stenocephala, Toxocara canis, Toxascaris leonina, or Trichuris vulpis) were enrolled in Group 1 (G1) and twenty healthy adult dogs were included in Group 2 (G2), which served as the control. A clinical assessment was performed using a physician-based scoring system. Oxidative stress variables and high-mobility group box 1 were assessed and compared by the means of unpaired t-tests (p < 0.05). Spearman's rank correlation was performed to calculate the correlation between oxidative stress variables, high-mobility group box 1, hematological parameters, and clinical signs. The results showed statistically significant values for reactive oxidative metabolites, thiol groups of plasma compounds, and high-mobility group box 1 in G1. Negative correlations between thiol groups and the number of red cells and hemoglobin were recorded. These preliminary results support the potential role of oxidative stress and HGMB-1 in the pathogenesis of gastrointestinal helminthiasis in dogs.

Differential Redox State and Iron Regulation in Chronic Obstructive Pulmonary Disease, Acute Respiratory Distress Syndrome and Coronavirus Disease 2019.

In patients affected by Acute Respiratory Distress Syndrome (ARDS), Chronic Obstructive Pulmonary Disease (COPD) and Coronavirus Disease 2019 (COVID-19), unclear mechanisms negatively interfere with the hematopoietic response to hypoxia. Although stimulated by physiological hypoxia, pulmonary hypoxic patients usually develop anemia, which may ultimately complicate the outcome. To characterize this non-adaptive response, we dissected the interplay among the redox state, iron regulation, and inflammation in patients challenged by either acute (ARDS and COVID-19) or chronic (COPD) hypoxia. To this purpose, we evaluated a panel of redox state biomarkers that may integrate the routine iron metabolism assays to monitor the patients' inflammatory and oxidative state. We measured redox and hematopoietic regulators in 20 ARDS patients, 20 ambulatory COPD patients, 9 COVID-19 ARDS-like patients, and 10 age-matched non-hypoxic healthy volunteers (controls). All the examined pathological conditions induced hypoxia, with ARDS and COVID-19 depressing the hematopoietic response without remarkable effects on erythropoietin. Free iron was higher than the controls in all patients, with higher levels of hepcidin and soluble transferrin receptor in ARDS and COVID-19. All markers of the redox state and antioxidant barrier were overexpressed in ARDS and COVID-19. However, glutathionyl hemoglobin, a candidate marker for the redox imbalance, was especially low in ARDS, despite depressed levels of glutathione being present in all patients. Although iron regulation was dysfunctional in all groups, the depressed antioxidant barrier in ARDS, and to a lesser extent in COVID-19, might induce greater inflammatory responses with consequent anemia.