Oxidative Modification Of Proteins Research Articles

High–intensity exercise training induces the oxidative modification of malate dehydrogenase 2 in skeletal muscles

High–intensity exercise training abrogates the aerobic metabolic system in skeletal muscles. Mitochondria play a crucial role in aerobic metabolism and regulate energy supply for muscle contraction. Although the detailed mechanism of mitochondrial dysfunction remains unknown, excessive reactive oxygen species (ROS) generated in response to high–intensity exercise may be involved. Herein, we examined the oxidative modification of mitochondrial proteins in mouse muscle following high–intensity exercise training. Male ICR mice (10–week–old) were divided into sedentary and high–intensity exercise groups. Mice in the exercise group received treadmill training five times per week for two weeks. The levels of oxidative protein modifications and the factors related to mitochondrial biogenesis and dynamics in the gastrocnemius muscle were measured. The levels of hexanoyl lysine adduct (HEL) and 4–hydroxy–2–nonenal (HNE) modified proteins tended to be higher in the muscle mitochondrial fraction, but not whole extractions, of the exercise group than in that of the sedentary group. Specifically, HEL and HNE modifications of malate dehydrogenase 2 (MDH2), a citric acid cycle–related enzyme, were particularly higher in the exercise group mice. Although mitochondrial biogenesis factors were upregulated by exercise, higher phosphorylation of dynamin–related protein 1 (Ser637) was observed in the exercise group. These results suggest that oxidative modification of MDH2 impairs the metabolic system following high–intensity exercise training, which may be associated with attenuated adaptation.

Beneficial Effects of Two Hydrogen Sulfide (H2S)-Releasing Derivatives of Dexamethasone with Antioxidant Activity on Atopic Dermatitis in Mice.

Hydrogen sulfide (H2S) is particularly produced in the skin, where it participates in the regulation of inflammation, pruritus, cytoprotection, scarring, and angiogenesis. In this study, we compared the effects of dexamethasone (Dex) with two H2S-releasing Dex derivatives in a murine model of atopic dermatitis (AD) induced by topical application of 2,4-dinitrochlorobenzene (DNCB). After sensitization with DNCB, the animals were topically treated for five consecutive days with either the H2S-releasing compounds 4-hydroxy-thiobenzamide (TBZ) and 5-(p-hydroxyphenyl)-1,2-dithione-3-thione (ADT-OH), Dex, or the derivatives Dex-TBZ or Dex-ADT. Topical treatment with equimolar doses of either Dex, Dex-TBZ, or Dex-ADT resulted in similar reductions in dermatitis score, scratching behavior, edema, eosinophilia, splenomegaly, and histological changes. In contrast with Dex, the H2S-releasing derivatives prevented IL-4 elevation and oxidative modification of skin proteins. On an equimolar dose basis, Dex-TBZ, but not Dex-ADT, promoted the elevation of endogenous H2S production and GPx activity. Neither Dex-TBZ nor Dex-ADT decreased GR activity or caused hyperglycemia, as observed with Dex treatment. We conclude that the presence of H2S-releasing moieties in the Dex structure does not interfere with the anti-inflammatory effects of this corticosteroid and adds beneficial therapeutical actions to the parent compound.

Anti-inflammatory and antioxidant actions of extracts from Rheum rhaponticum and Rheum rhabarbarum in human blood plasma and cells in vitro

Rheum rhaponticum L. (rhapontic rhubarb) and Rheum rhabarbarum L. (garden rhubarb) are edible and medicinal rhubarb species used for many centuries in traditional medicine. This work is focused on the biological activity of extracts from petioles and roots of R. rhaponticum and R. rhabarbarum as well as rhapontigenin and rhaponticin, typical stilbenes present in these rhubarbs, in a context of their effects on blood physiology and cardiovascular health. Anti-inflammatory properties of the examined substances were evaluated in human peripheral blood mononuclear cells (PBMCs) and THP1-ASC-GFP inflammasome reporter cells. Due to the coexistence of inflammation and oxidative stress in cardiovascular diseases, the study design included also antioxidant assays. This part of the work involved the assessment of the protective efficiency of the examined substances against the peroxynitrite-triggered damage to human blood plasma components, including fibrinogen, a protein of critical importance for blood clotting and maintaining the haemostatic balance. Pre-incubation of PBMCs with the examined substances (1–50 μg/mL) considerably decreased the synthesis of prostaglandin E2 as well as the release of pro-inflammatory cytokines (IL-2 and TNF-α) and metalloproteinase-9. A reduced level of secreted apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) specks in the THP-1-ASC-GFP cells was also observed. The examined substances significantly diminished the extent of ONOO‾induced oxidative modifications of blood plasma proteins and lipids and normalized, or even strengthened blood plasma antioxidant capacity. Furthermore, a reduction of oxidative damage to fibrinogen, including modifications of tyrosine and tryptophan residues along with the formation of protein aggregates was found.

Oxidative homeostasis in germinating pea seeds (isum sativum) depending on ultrasonic exposure duration

The effects of ultrasonic exposure durations (5, 10 and 20 min) with intensity of 25 kW/m2 and the frequency of 26.1 kHz on the extent of lipid peroxidation, oxidative modification of proteins, the activity of cysteine proteinases, and gene expression in germinating pea seeds were studied. It has been shown that the intensity in the area that had seeds planted (just over the central ultrasound generator) was rather uniformly distributed, but at the same time gave rise to diffuse reflection. A range of tendencies in changes of the indicators under study were seen after ultrasound exposure. The level of malonic dialdehyde increased with increasing duration of ultrasound exposure. This study revealed the increased amount of oxidized proteins in germinating pea seeds after a 10 min-exposure to ultrasound, and a decrease in the level of oxidative modification of proteins present in seeds subjected to ultrasound for a period of 20 min. The activity of cysteine proteinases was higher in pea seeds after a 5-min exposure to ultrasound, but the amount of mRNA transcripts increased in all experimental samples.

Significance of Type II Collagen Posttranslational Modifications: From Autoantigenesis to Improved Diagnosis and Treatment of Rheumatoid Arthritis.

Collagen type II (COL2), the main structural protein of hyaline cartilage, is considerably affected by autoimmune responses associated with the pathogenesis of rheumatoid arthritis (RA). Posttranslational modifications (PTMs) play a significant role in the formation of the COL2 molecule and supramolecular fibril organization, and thus, support COL2 function, which is crucial for normal cartilage structure and physiology. Conversely, the specific PTMs of the protein (carbamylation, glycosylation, citrullination, oxidative modifications and others) have been implicated in RA autoimmunity. The discovery of the anti-citrullinated protein response in RA, which includes anti-citrullinated COL2 reactivity, has led to the development of improved diagnostic assays and classification criteria for the disease. The induction of immunological tolerance using modified COL2 peptides has been highlighted as a potentially effective strategy for RA therapy. Therefore, the aim of this review is to summarize the recent knowledge on COL2 posttranslational modifications with relevance to RA pathophysiology, diagnosis and treatment. The significance of COL2 PTMs as a source of neo-antigens that activate immunity leading to or sustaining RA autoimmunity is discussed.

Effect of Low-Dose Line-Spectrum and Full-Spectrum UV on Major Humoral Components of Human Blood.

Ultraviolet blood irradiation (UVBI) is an alternative approach to the treatment of infectious diseases of various pathogeneses. Recently, UVBI has attracted particular interest as a new immunomodulatory method. Experimental studies available in the literature demonstrate the absence of precise mechanisms of the effect of ultraviolet radiation (UV) on blood. Here, we investigated the effect of UV radiation of line-spectrum mercury lamp (doses up to 500 mJ/cm2) traditionally used in UVBI on the major humoral blood components: albumin, globulins and uric acid. Preliminary data on the effect of various doses of UV radiation of full-spectrum flash xenon lamp (doses up to 136 mJ/cm2), a new promising source for UVBI, on the major blood plasma protein, albumin, are presented. The research methodology included spectrofluorimetric analysis of the oxidative modification of proteins and analysis of the antioxidant activity of humoral blood components by chemiluminometry. The effect of UV radiation on albumin caused its oxidative modification and, accordingly, an impairment of the transport properties of the protein. At the same time, UV-modified albumin and γ-globulins acquired pronounced antioxidant properties compared to native samples. Uric acid mixed with albumin did not protect the protein against UV-induced oxidation. The flash full-spectrum UV qualitatively had the same effect on albumin as line-spectrum UV did, but an order of magnitude lower doses were required to achieve comparable effects. The suggested protocol can be used for selecting a safe individual dose for UV therapy.

EFFECTIVENESS AND SAFETY OF CHONDROGUARD THERAPY IN PATIENTS WITH ENDOCRINE PHENOTYPE OF OSTEOARTHRITIS

Objective. To evaluate the efficiency and safety of therapy with parenteral chondroitin sulfate (CS) Chondroguard®, CS-Bioactive© (Bioiberica S.A.U., Spain); ZAO “Sotex” PharmFirm, Russia) in patients with Stage II knee osteoarthritis (KOA), grade 1 knee functional insufficiency (FI), and type 2 diabetes mellitus (T2DM). Subjects and methods. A 50-day comparative pilot study involved 80 patients with Kellgren-Lawrence Stage II KOA, grade 1 FI, and T2DM, who were divided into 2 groups; Group 1 (a study group; n=40) received CS (Chondroguard®, 100 mg/ml, intramuscularly, every other day; the first three injections of 1 ml each; if they were well tolerated; starting from injection 4 of ml each; the treatment cycle was 25 injections for 50 days) and therapy for T2DM and its complications; Group 2 (a comparison group; n=40) took oral symptomatic slow acting drugs for osteoarthritis (SYSADOA) agents containing CS, glucosamine, unsaponifiable compounds of avocado and soybean, and therapy for T2DM and its complications. On study days 0 and 50, the investigators calculated in the patients the scores of the visual analogue scale (VAS), WOMAC index, the Lequesne index, those of the WHO and the Narango Therapy Safety Scales, assessed the parameters of the blood hormonal and metabolic profile, free radical oxidation (FRO), antioxidant system (AOS), and the levels of blood proinflammatory factors. Results. In Group 1, therapy with CS was well tolerated and was accompanied by a reduction in pain intensity on VAS at rest and in motion, in functional restrictions on the WOMAS index and the Lequesne index, as well аs by a decrease in blood proinflammatory markers (С-reactive protein, interleukin 6 and 1β, leptin, lipocalin-2), in the concentration of fasting and postprandial glucose, HbA1c, С-peptide, insulin-like growth factor-1, and in FRO-AOS activity (malondialdehyde, protein oxidative modification, superoxide dismutase) as compared with the indicators in Group 2. Conclusion. Chondroguard® was shown to be effective and safe in the combination therapy in patients with Stage II endocrine KOA phenotype, grade 1 FI, and T2DM.

Cysteine carbonylation with reactive carbonyl species from lipid peroxidation induce local structural changes on thioredoxin active site

Protein oxidative modifications with reactive carbonyl species (RCS) is directly linked to metabolic processes in premature aging, cancer, neurodegenerative and infectious diseases. RCS as 4-Hydroxy-2-nonal (HNE), 4-Hydroxy-2-hexenal (HHE), 4-Oxo-2-nonenal (ONE) and Malondialdehyde (MDA) attack nucleophilic amino acids residues forming irreversible adducts with proteins as Thioredoxins (Trx). This is a class of small thiol oxide-reductases playing a key role in redox signaling and oxidative stress responses in mammals. Although proteomic studies have identified to Cys-32 residue as a target of HNE attack that inhibit its enzymatic activity, how this carbonylation affects its structure and dynamic behavior at the atomic level is unknown. Even more, the molecular bases for the atomistic behavior of these modified proteins have not been completely understood. We present molecular dynamics simulations of Trx-modified with four different RCS to analyze its global and local structural effects. For this, parameters supported in the AMBER force fields were built and validated for three non-natural cysteine residues modified with HHE, ONE and MDA. Results obtained showed a slight change in the global conformational stability of Trx modified with HNE and MDA, establishing that all modified proteins presented local regions of high mobility in the modified catalytic site and some regions far from the modification area. In addition, essential domain movement modes evidences that proteins modified with the RCS assayed induce changes in conformational flexibility. Finally, these data showed that the given conformational changes did not caused global changes in proteins but rather localized changes in particular regions.

Analysis of clinical and paraclinical findings in children with community-acquired pneumonia and comorbid chronic tonsillitis

Background. Respiratory morbidity is the most frequent cause for children consulting a doctor, accounting for about one-quarter of primary care consultations. The current research aimed to analyze the clinical and paraclinical features of community-acquired pneumonia (CAP) with comorbid chronic tonsillitis in children and to assess the prognostic value of the proposed diagnostic procedures to optimize the management of patients. Materials and methods. The study was conducted at the Pulmonology and Allergology and Infectious Department of the Municipal Medical Establishment “Chernivtsi Regional Children’s Clinical Hospital”, Ukraine. Clinical group I included 20 children (mean age 9.40 ± 1.56 years) with uncomplicated community-acquired pneumonia and concomitant chronic tonsillitis, and the clinical group II consisted of 16 patients (mean age 9.60 ± 1.13 years) with CAP without concomitant pathology of the upper respiratory tract. Results. Community-acquired pneumonia in children with comorbid chronic tonsillitis is characterized by an intense proinflammatory response in the airways, which manifests itself in fever (odds ratio (OR) 6.0), malaise and fatigue (OR 14.3), persistent cough and dyspnea (OR 3.7), leukocytosis (OR 2.0), high levels of acute phase proteins (OR 4.5), activation of the oxidative protein modification according to the exhaled breath condensate examination (OR 6.0–30.0), and more frequent fungal-bacterial association according to the microbiological examination of sputum and oropharyngeal swab (OR 11.1). The obtained data suggest that parenteral antibiotic therapy was used in 20 % of patients from group I and 12.5 % from group II, while 6.3 % of children from group II received only oral antibiotics. In children with CAP and chronic tonsillitis versus comparison group, the ОR of receiving parenteral antibiotic therapy for longer than 7 days reached 10.0, and the OR of oral antibiotic therapy for longer than 5 days reached 4.0. Conclusions. The results obtained from the study show that children with CAP and comorbid chronic tonsillitis have more pronounced clinical symptoms, accompanied by paraclinical signs of inflammation that last longer during hospital treatment and require comprehensive therapy (OR 3.3) of longer duration (OR 4.0–10.0).

IN VITRO ANTIOXIDANT ACTIVITY OF COMPOSITIONS BASED ON BIOSURFACTANTS AND AMINO-CONTAINING HETEROCYCLIC DERIVATIVES OF 1,4-NAPHTOQUINONE

Compositions of rhamnolipids (RL) and trehalosе lipids (TL) with amino-containing heterocyclic derivatives of 1,4-naphthoquinone were obtained for the first time. The acute toxicity of amino-containing derivatives of 1,4-naphthoquinone was determined by in silico method using the GUSAR program. We have studied influence of selected compounds and their rhamnolipid and trehalosе lipids compositions on the processes of lipid peroxidation (LPO) and oxidative modification of proteins (OMP) in rat liver tissues upon initiation of free radical oxidation in vitro. It was determined that 1,4-naphthoquinone derivatives have antioxidant activity. RL and TL have a pro-oxidant effect, increase the content of secondary products of lipoperoxidation compared to the control. They have an antioxidant effect in a composition with 1,4-naphthoquinone derivatives. The lead-compound 2-[(6-(4-fluorophenyl-5-oxo-2,5-dihydro-1,2,4-triazin-3-yl)phenyl)amino]naphthalene-1,4-dione (1) and its complex with rhamnolipid show high antioxidant activity (which is confirmed by the decrease in the content of carbonyl groups in the side chains of proteins compared to the control).

Identification of autoantigens and their potential post-translational modification in EGPA and severe eosinophilic asthma.

The chronic airway inflammation in severe eosinophilic asthma (SEA) suggests potential autoimmune aetiology with unidentified autoantibodies analogous to myeloperoxidase (MPO) in ANCA-positive EGPA (eosinophilic granulomatosis with polyangiitis). Previous research has shown that oxidative post-translational modification (oxPTM) of proteins is an important mechanism by which autoantibody responses may escape immune tolerance. Autoantibodies to oxPTM autoantigens in SEA have not previously been studied. Patients with EGPA and SEA were recruited as well as healthy control participants. Autoantigen agnostic approach: Participant serum was incubated with slides of unstimulated and PMA-stimulated neutrophils and eosinophils, and autoantibodies to granulocytes were identified by immunofluorescence with anti-human IgG FITC antibody. Target autoantigen approach: Candidate proteins were identified from previous literature and FANTOM5 gene set analysis for eosinophil expressed proteins. Serum IgG autoantibodies to these proteins, in native and oxPTM form, were detected by indirect ELISA. Immunofluorescence studies showed that serum from patients with known ANCA stained for IgG against neutrophils as expected. In addition, serum from 9 of 17 tested SEA patients stained for IgG to PMA-stimulated neutrophils undergoing NETosis. Immunofluorescent staining of eosinophil slides was evident with serum from all participants (healthy and with eosinophilic disease) with diffuse cytoplasmic staining except for one SEA individual in whom subtle nuclear staining was evident. FANTOM5 gene set analysis identified TREM1 (triggering receptor expressed on myeloid cells 1) and IL-1 receptor 2 (IL1R2) as eosinophil-specific targets to test for autoantibody responses in addition to MPO, eosinophil peroxidase (EPX), and Collagen-V identified from previous literature. Indirect ELISAs found high concentrations of serum autoantibodies to Collagen-V, MPO, and TREM1 in a higher proportion of SEA patients than healthy controls. High concentrations of serum autoantibodies to EPX were evident in serum from both healthy and SEA participants. The proportion of patients with positive autoantibody ELISAs was not increased when examining oxPTM compared to native proteins. Although none of the target proteins studied showed high sensitivity for SEA, the high proportion of patients positive for at least one serum autoantibody shows the potential of more research on autoantibody serology to improve diagnostic testing for severe asthma. ClinicalTrials.gov, identifier, NCT04671446.

RESTRICTED PROTEOLIS AND OXIDIZING MODIFICATION OF PROTEINS IN HEPATOCYATES OF PATIENTS WITH DIABETES AND TUBERCULOSIS

Objective: The objective of the work is to investigate the oxidative modification of proteins and limited proteolysis in hepatocytes in patients with diabetes and pulmonary tuberculosis. Design and method: A prospective pathomorphological study of 60 cases of death of patients who died of various causes, in which in the final clinical and pathologoanatomical diagnosis as the main disease appeared pulmonary tuberculosis. Depending on clinical forms and variants of tuberculosis, the main group was divided into three subgroups. Results: An important role in the pathogenesis of the formation of both diabetes mellitus and tuberculosis play an impairment of immune homeostasis, especially in patients with expressive symptoms of exacerbation of the disease and prolonged relapse. Oxidative stress leads to an increase in the production of active forms of oxygen and as a consequence - oxidation modification of proteins (OMB): their aggregation and fragmentation, which is accompanied by irreversible changes in the physical, chemical and biological properties of the protein molecule. Taking into account the different functional activity of hepatocytes of different zones of the acinus (according to Rappoport) according to their own research, the R / B coefficient (specific coloration on acid and main proteins for Michel Calvo) and average values of relative units of optical density of a specific color on the free amino group of proteins for A. Yasuma and T. Ichikava were determined separately in the hepatocytes of the first, second and third zones and established the linear nature of the increase in the intensity of the oxidative modification of proteins and limited proteolysis from the 1st to the 3rd zone (from peripartal to centrolobular branches of acinus) in patients with resistant forms of tuberculosis. Conclusions: Diabetes mellitus in patients with lung tuberculosis leads to increased processes of free radical oxidation of proteins with characteristic effects - increased proteolysis and oxidation of amino groups of proteins – in hepatocytes of the liver.

Study of the immunoglobulin and oxidized protein content of semen under infertility

Among the causes of male infertility, enough attention is paid to oxidative stress, which in turn is a pathogenetic link in the inflammatory process. However, there is practically no information on the content of oxidized modified proteins in the semen, which makes it difficult to study the pathogenesis of diseases of the male reproductive system. In part, protein oxidation may be due to the production of reactive oxygen species by microorganisms, both directly and indirectly through the activation of immune system cells. The aim of the research was to study the level of oxidized modified proteins and changes in immunoglobulin concentrations in the semen under bacteriospermia. A study was made of the ejaculate of 48 men who applied to the clinic for infertility in marriage. The comparison group consisted of 32 practically healthy men who had no growth of microorganisms in the ejaculate samples. When conducting bacteriological analysis, the studied samples were diluted 10 times and used the generally accepted method. The concentration of albumin, immunoglobulins A, M, G, E was determined in the spermatic fluid. The oxidative modification of proteins was evaluated in the reaction with 2,4-dinitrophenylhydrazine. The concentration of oxidized proteins was expressed in nmol/mg of the total protein of the studied biological fluid. The biuret method was used to determine the protein concentration. Statistical analysis of the results was performed using descriptive statistics and Student's t-test for paired data. The concentration of protein in the seminal fluid did not differ significantly among the studied groups. The albumin concentration (16.96±1.28 mg/mL) was statistically significantly lower in the absence of microorganism growth than in bacteriospermia. With bacteriospermia, a decrease in the concentration of IgM and IgA and an increase in the level of IgG were noted. The degree of protein oxidation is maximum when enterobacteria are isolated from seminal fluid. Thus, during the studies it was found that, despite the absence of a clinic, with asymptomatic bacteriospermia, the secretion of immunoglobulins G into the semen is observed. The accumulation of oxidized proteins in the seminal fluid in bacteriospermia has been shown.

Detection of Cysteine Sulfenic Acid on E. coli Proteins with a Biotin-Benzoboroxole Probe.

S-sulfenylation of cysteine residues on proteins can effectively change protein structures and accordingly regulate their functions in vivo. Investigation of S-sulfenylation in different biological environments is thus vital for a systematic understanding of cellular redox regulation. In this work, a functional probe, biotin-benzoboroxole (Bio-ben), was designed for the detection of cysteine sulfenic acid (Cys-SOH). The performance of Bio-ben was characterized by small-molecule sulfenic acid, protein models, and proteome tests via mass spectra and western blotting. The results showed that Bio-ben was validated for cysteine sulfenic acid on proteins with good capture efficiency even at low concentrations. Compared with commonly used probes such as dimedone, the current probe has significantly shortened labeling time and exhibited comparable sensitivity. The proposed method provides a new approach for exploring S-sulfenylation in the oxidative modification of proteins and is helpful for related biological and clinical applications.

OXIDATIVE MODIFICATION OF PROTEIN AND RESERVE-ADAPTIVE POTENTIAL IN PATIENTS WITH METABOLIC PHENOTYPE OF OSTEOARTHRITIS

Relevance. Oxidative stress, resulting from an imbalance between ROS production and the antioxidant defense system, is a major disruption of chronic inflammation in OA and obesity. It should be noted that the study and understanding of OS and their relationship with the identification of joint tissues is necessary to develop new therapeutic approaches to the occurrence and expediency of the metabolic phenotype of OA. Aim. To study protein oxidative modification (OMP) and reserve-adaptive potential in patients with the metabolic phenotype of osteoarthritis (OA). Material and methods. Participants were divided into 2 groups: Control group - patients without articular pathology, metabolic syndrome. The exper-imental group consisted of patients with the metabolic phenotype of OA. The subjects were collected complaints and anamnesis, as well as general clinical and orthopedic examination. In the blood serum, indicators of OMP and superoxidedismutase (SOD) were determined. Patients were asked to answer the questions of clinical scales of the functional state of the joints and quality of life. Results. The initial activity of blood serum SOD was somewhat lower in patients with the metabolic phenotype of OA, and the level of OMP showed certain differences in the levels of activity of the processes of spontaneous and metal-catalyzed PMB. The total area under the spontaneous OMP curve was statistically significantly higher (p 0,01), mainly due to the neutral fraction of aldehyde dinitrophenyl hydrozones (ADNFH). The level of reserve-adaptive potential was statistically significantly lower than the control group. An inverse correlation was found between the total area, the area of ADNFH of metal-catalyzed OMP and the level of pain, which indicates higher levels of oxidative stress in patients with severe clinical symptoms. Сonclusion. In patients with pronounced indicators of clinical manifestations of OA, more active processes of redox changes were observed. There was a decrease in the activity of antioxidant activity and the level of reserve-adaptive potential

Influence of liposomal thiosulfonate drug on the blood parameters of cows suffering catarrhal mastitis

Diseases of the mammary gland in cows are of multi-factor nature, which significantly complicates their control and treatment. One of the factors influencing development of mastitis is bacterial microflora. Because of limited use of antibiotics, development of novel alternative antmicrobial drugs is of great relevance. Therefore, the objective of the study was identifying the effect of an ethyl-thiosulfanilate-based liposomal drug on the hematological and biochemical parameters of blood of the cows suffering catarrhal mastitis. The animals were three times intramuscularly injected with the liposomal drug in the dose of 0.04 mL/kg of body weight. Blood from the cows was taken from the jugular vein on the day prior to administration of the drug, and also on the 3rd and 7th days after its application. During the bacterial study of the secretion of udder of sick cows, we collected 51 bacterial isolates, in which dominated Staphylococcus aureus (27.5%) and Streptococcus spp. (21.6%). In blood of the sick cows, we found higher number of leukocytes, increases in the concentrations of circulating immune complexes, lipid hydroperoxides, and ketone derivatives of oxidative modification of proteins. At the same time, we observed decreases in the number of lymphocytes and concentration of protein and the bactericidal activity of blood serum. Administration of the complex liposomal drug had a normalizing effect on the analyzed parameters of the cows. This was evidenced by 9.4% decrease in the total number of leukocytes, 18.5% decrease in the content of circulating immune complexes, 9.3% increase in lymphocytes, and 13.6% increase in the level of total protein. In the sick cows, the drug enhanced the bactericidal (by 35.4%) and lysozymic (by 36.3%) activities of blood serum, glutathione peroxidase activity, and increased the content of reduced glutathione. On the seventh day after injecting the drug into blood of the cows, we saw decrease in the level of intermediate and end products of lipid peroxidation and derivatives of oxidative modification of proteins. Therefore, the complex ethyl-thiosulfanilate-based liposomal drug promoted reduction of metabolic homeostasis of the organism, had positive effect on the activity of natural defense mechanisms in the organism and can be recommended for the treatment of cows with catarrhal mastitis.

Human sperm proteome reveals the effect of environmental borne seminal polyaromatic hydrocarbons exposome in etiology of idiopathic male factor infertility.

Introduction: Polyaromatic hydrocarbons (PAHs) are considered as redox active environmental toxicants inducing oxidative stress (OS) mediated injury to cells. Oxidative predominance is reported in 30%-80% of idiopathic male infertility (IMI) patients. Hence, this work aims to unravel correlation, if any, between seminal PAH exposome and sperm function in IMI patients through a proteomic approach. Methods: Seminal PAH exposome was analyzed in 43 fertile donors and 60 IMI patients by HPLC and receiver operating characteristic (ROC) curve was applied to find out the cut-off limits. Spermatozoa proteome was analyzed by label free liquid chromatography mass spectroscopy (LC-MS/MS) followed by molecular pathway analysis using bioinformatic tools. Validation of key proteins' expression and protein oxidative modifications were analyzed by western blot. Results and discussion: Of the 16 standards toxic PAH, 13 were detected in semen. Impact of the different PAHs on fertility are Anthracene < benzo (a) pyrene < benzo [b] fluoranthene < Fluoranthene < benzo (a) anthracene <indol (123CD) pyrene < pyrene < naphthalene < dibenzo (AH) anthracene < fluorene < 2bromonaphthalene < chrysene < benzo (GH1) perylene as revealed by ROC Curve analysis (AUCROC). Benzo [a] pyrene is invariably present in all infertile patients while naphthalene is present in both groups. Of the total 773 detected proteins (Control: 631 and PAH: 717); 71 were differentially expressed (13 underexpressed, 58 overexpressed) in IMI patients. Enrichment analysis revealed them to be involved in mitochondrial dysfunction and oxidative phosphorylation, DNA damage, Aryl hydrocarbon receptor (AHR) signaling, xenobiotic metabolism and induction of NRF-2 mediated OS response. Increased 4-hydroxynonenal and nitrosylated protein adduct formation, and declined antioxidant defense validates induction of OS. Increased GSH/GSSG ratio in patients may be an adaptive response for PAH metabolism via conjugation as evidenced by over-expression of AHR and Heat shock protein 90 beta (HSP90β) in patients. Seminal PAH concentrations, particularly benzo (a) pyrene can be used as a marker to distinguish IMI from fertile ones with 66.67% sensitivity and 100% specificity (95% confidence interval) along with oxidative protein modification and expression of AHR and HSP90β.

Oxidative modification of proteins and antioxidant status in blood of the rats with experimental acute generalized peritonitis against the background of streptozotocin-induced diabetes

Modern aspects of the pathogenesis of acute inflammation of the peritoneum that is concurrent with diabetes involves analysis of metabolic mechanisms, in particular peroxidaton of proteins – antioxidant defense. Therefore, the objective of our study was to examine the interrelation between the processes of free-radical oxidation of proteins and antioxidant system in the dynamics of development of acute generalized peritonitis against the background of streptozotocin-induced diabetes. The study was performed on 56 non-linear white mature male rats. Diabetes mellitus was modeled by a single intraperitoneal injection of streptozotocin (60 mg/kg). On the 14th day of the development of streptozotocin-induced diabetes, we injected 10% filtrated faeces suspension (0.5 mL) into the abdominal cavity, thus initiating acute generalized peritonitis. Oxidative modification of proteins in blood serum was studied using the I. F. Meshchyshen’s method and the condition of antioxidant protection was monitored according to the activities of superoxide dismutase, catalase, content of reduced glutathione, and the level of ceruloplasmin. The study of the parameters of free-radical oxidation of proteins and study of the condition of antioxidant system in blood of the rats with experimental acute generalized peritonitis against the background of streptozotocin-induced diabetes demonstrated that oxidative protein modification grew, while the parameters of the activity of antioxidant system were being inhibited, depending on the stage of acute inflammation of the peritoneum. We determined inverse correlation relationships between the products of free-radical oxidation of proteins and parameters of antioxidant system on the third and on the seventh days of modelling of combined pathology. Manifestation of acute generalized peritonitis concurring with streptozotocin-induced diabetes was accompanied by a gradual accumulation of the products of free-radical oxidation of proteins and exhaustion of the antioxidant defense during all stages of the development of acute inflammation of the peritoneum, peaking on the seventh day after administration of faecal suspension (terminal stage of peritonitis). The observed inverse correlations between the levels of oxidative modification of proteins and the activity of superoxide dismutase, catalase, reduced glutathione, and ceruloplasmin on the third and on the seventh days of modelling of combined pathology indicate a predictive role of the processes of free-radical oxidation of proteins in exhaustion of antioxidant-defense resources.

Invitro impact of a combination of red and infrared LEDs, infrared laser and magnetic field on biomarkers of oxidative stress and hemolysis of erythrocytes sampled from healthy individuals and diabetes patients.

Low-intensity infrared laser irradiation with output emissions of the laser and LED for in vitro irradiation of plasma and erythrocyte samples collected from healthy individuals and diabetes mellitus (DM) patients was used in the current study. The generated emission was in the range 0.85-0.89nm with pulse duration near 130ns and repetition rates of pulses 50, 150, 600, and 1500Hz, average power 0, 50, or 100mW, in the range of 1-9min for different 30 variants of irradiation. The levels of 2-thiobarbituric-acid reactive substances (TBARS), aldehydic and ketonic derivatives of oxidatively modified proteins (OMP), total antioxidant capacity (TAC), acid-induced resistance of erythrocytes, and activities of the main antioxidant enzymes were assessed in erythrocyte and plasma samples after irradiation. The low-intensity infrared laser irradiation and low-intensity light emitted by a red LED decreased the lipid peroxidation levels in the erythrocytes of both healthy individuals and DM patients. A statistically significant decrease in TBARS and OMP levels and an increase in the TAC level were observed at the irradiation energy of 34.39 and 68.79J/cm2 for samples collected from both healthy individuals and DM patients. The effects of the irradiation were accompanied by a statistically significant decrease in catalase activity of both healthy individuals and DM patients. In many variants of the laser irradiation and low-intensity light emitted by a red LED used in our study, a decrease in the percent of hemolyzed erythrocytes was observed, suggesting that laser therapy protocols should take into account fluencies, frequencies, and wavelengths of the laser before the beginning of treatment, especially in DM patients.

Blood Plasma Calorimetric Profiles of Women with Preeclampsia: Effect of Oxidative Stress.

Preeclampsia is a pregnancy-related disease with poor placentation and presents itself through hypertension and proteinuria. The disease is also associated with the oxidative modification of proteins in maternal blood plasma. In this work, we combine differential scanning calorimetry (DSC), capillary electrophoresis, and atomic force microscopy (AFM) to evaluate the changes in the plasma denaturation profiles of patients with preeclampsia (PE) as compared with those of pregnant controls. Our results demonstrate that the last trimester of pregnancy substantially affects the main calorimetric characteristics of blood plasma from pregnant controls relative to nonpregnant women. These variations correlate well with the changes in protein levels determined by electrophoresis. DSC analysis revealed significant deviations in the plasma heat capacity profiles of preeclamptic patients from those of pregnant controls. These alterations are expressed mainly in a substantial reduction in albumin-assigned transitions and an upward shift in its denaturation temperature, lower calorimetric enthalpy changes, and a reduced ratio of heat capacity in the albumin/globulin-assigned thermal transitions, which are more pronounced in severe PE cases. The in vitro oxidation model shows that the alteration of PE thermograms is partly related to protein oxidation. AFM data detected numerous aggregate formations in the plasma of PE samples and fewer small ones in the pregnant controls, which are not found in healthy nonpregnant samples. These findings could serve as a basis for further investigations to reveal the possible relationship between albumin thermal stabilization, the increased inflammatory state and oxidative stress, and protein misfolding in preeclampsia.