[Serum brain damage biomarkers as a diagnostic and prognostic tool in ischemic stroke]. / Syvorotochnye biomarkery porazheniya mozga kak diagnosticheskii i prognosticheskii instrument pri ishemicheskom insul'te.

OBJECTIVE: To study serum quantities of neuron specific enolase (NSE), glial fibrillary acidic protein (GFAP) and NR2-antibodies (NR2-ab) in various cerebrovascular pathology and assess their value as a panel used as a diagnostic and predictive tool for stroke. MATERIAL AND METHODS: NSE, GFAP and NR2-ab serum levels were measured twice for 84 patients with ischemic stroke (IS) and 8 patients with hemorrhagic stroke (HI), once for 8 patients with transient ischemic attack (TIA), 26 patients with chronic brain ischemia (CBI), 27 healthy volunteers (HV). RESULTS: NSE and GFAP levels were significantly higher in IS than in CBI and HV patients, and NR2-ab levels in IS were higher than in TIA and lower than in HV. In patients with more pronounced neurological deficiency and less favorable functional outcome by day 10-14 of IS, the levels of NSE, GFAP and NR2-ab were higher. Sensitivity and specificity of biomarker panel was higher than with their separate application. CONCLUSION: The NSE, GFAP and NR2-ab biomarkers have a diagnostic and predictive value for IS.

The state of platelet and plasma hemostasis as a predictor of coronary blood flow in patients with acute myocardial infarction.

Aim      To study the relationship of the platelet function and plasma homeostasis with the blood flow in the infarct-related artery (IRA) and with the course of acute myocardial infarction (AMI).Material and methods  This study included 93 patients with AMI (75 patients with ST-elevation AMI and 18 patients without ST segment elevation). 63 patients had TIMI 0-1 blood flow in the IRA and 30 patients had TIMI 2-3. Rotational thromboelastometry, impedance aggregometry, the endothelium-dependent vasodilation (EDVD) test, and the thrombodynamics test were performed for all patients. The primary clinical endpoint included the totality of in-hospital complications of AMI, and the secondary endpoint included the totality of out-of-hospital complications of AMI. Major bleedings (BARC 3-5) and minor bleedings (BARC 1-2) were evaluated separately.Results Patients with IRA TIMI 0-1 flow were characterized by a shorter blood clotting time (BCT), larger thrombus size and density, more intense platelet aggregation induced by arachidonic acid and ADP, and lower values of the EDVD test. It was found that the parameters of platelet aggregation induced by arachidonic acid (AUC Asa) in combination with BCT allowed assessment of the severity of IRA blood flow disorder (sensitivity 76 %, specificity 71 %) in patients with AMI, regardless of the presence of ST segment elevation on the ECG. In addition, the incidence of the primary endpoint was greater in patients with IRA TIMI 0-1 flow (41.3% and 16.7%, respectively; p=0.015). In patients with TIMI 2-3 flow in the long-term period of the disease, the incidence of minor bleedings was significantly higher (8.5% and 30.4 %, respectively; p=0.045).Conclusion      Compared to patients with preserved blood flow, patients with AMI and IRA TIMI 0-1 flow are characterized by endothelial dysfunction and more intense processes of thrombogenesis and platelet aggregation. It has been shown for the first time that the combination of two simple criteria for assessing hemostasis (AUC Asa; BCT) allows assessment of the degree of IRA blood flow disorder in patients with AMI.

[The role of thrombin in the pathogenesis of atherosclerosis and its complications].

Thrombin is a key regulator of the homeostasis system. Also, it actively participates in progression of various systemic diseases, including atherosclerosis. There is a large amount of experimental and clinical data on the involvement of thrombin in the pathogenesis of ischemic heart disease (IHD). Thus, studying thrombin activity regulation is promising. Also, the question whether it is possible to use biomarkers of thrombin activity as predictors of cardiovascular complications in IHD patients is relevant. The present review focuses on major mechanisms of thrombin functioning, its role in development and progression of atherosclerosis, and available tests for evaluation of thrombin functional activity. Major clinical studies are discussed that evaluated the efficacy of thrombin inhibitors and protease-activated receptor antagonists.

Duration of Applications Affects the Long-Term Efficacy of Cryoballoon Ablation Pulmonary Veins.

Aim      To evaluate the effect of cryo-exposure duration and the use of the Achieve circular mapping catheter on efficacy of cryoballoon ablation (CBA).Material and methods  CBA of pulmonary vein ostia (PVO) is a major method for heart rhythm control in patients with atrial fibrillation (AF). Since the inception, the PVO CBA method has evolved; the recommended application time was changed, and the Achieve circular catheter appeared. We performed a retrospective analysis of PVO CBA administered to patients with AF in the I.V. Davydovsky Municipal Clinical Hospital from 2017 through 2019. The study included 100 patients with available clinical and demographic characteristics and remote results of the intervention. Three patient groups were analyzed based on differences in surgical techniques: group 1, Guidewire / 240 (n=31) with the cryoballoon placing on a guidewire and PVO exposure duration of 240 s; group 2, Guidewire / 180 (n=26) with the cryoballoon placing on a guidewire and PVO exposure duration of 180 s; and group 3, Achieve / 180 (n=43) with the cryoballoon placing on the mapping catheter Achieve and PVO exposure duration of 180 с. The follow-up period was 33.2±4.5, 15.2±6.1, and 12.2±4.1 months in the Guidewire / 240, Guidewire / 180, and Achieve / 180 groups, respectively. The intervention was considered effective when there was no relapse at the time of interview. A relapse of AF was determined as one or more paroxysms recorded on electrocardiogram (ECG) or during 24-h ECG monitoring; the "blind period" (first 3 months after the procedure) was excluded from the follow-up. Safety evaluation included clinically significant complications, such as phrenic nerve damage, hemopericardium, gastroparesis, hemoptysis, acute cerebrovascular disease, and formation of atrio-esophageal fistula. Effects of independent factors were determined with binary logistic regression.Results In the Guidewire / 240 group, efficacy of PVO CBA for the maximum follow-up period was 74.4%, which was significantly different from the value for the Guidewire / 180 group (57.7 %, р=0.015). At the same time, the difference between the Guidewire / 240 and Achieve / 180 groups was statistically non-significant for a comparable follow-up period (р=0.144). Clinically significant complications were absent in all 3 groups. The independent factors that significantly increased the PVO CBA efficacy were the cryo-exposure duration of 240 s compared to 180 s (р= 0.018) and the use of the Achieve catheter (р=0.014).Conclusion      Decreasing the cryo-exposure duration to less than 240 s is impractical (in absence of Achieve mapping catheter) since it impairs the long-term efficacy of PVO CBA and does not influence the risk of complications.

Cytomegalovirus Infection in Cardiovascular Diseases.

Atherosclerosis underlies the development of many cardiovascular diseases that continue to hold a leading place among the causes of death in developed countries. The role of activated immune cells in atherosclerosis progression has been convincingly demonstrated, but the mechanism of their action remains poorly investigated. Since atherosclerosis is associated with chronic inflammatory response, involvement of viral and bacterial infections in atherogenesis has been examined. A special place among the infectious agents is held by human herpesviruses as the most common persistent viruses in human population coupled to chronic inflammation during atherosclerosis. We found that activation of cytomegalovirus (CMV, human herpesvirus 5) infection is associated with the emergence of acute coronary syndrome, which is in a good agreement with the data on productive CMV infection published elsewhere. In this review, we discuss the data obtained by us and other researchers regarding the role of cytomegalovirus infection and related potential mechanisms resulting in the expansion of atherosclerotic plaques during ischemic heart disease and stroke, including virus transfer to immune and endothelial cells via extracellular vesicles. In particular, the data presented in the review demonstrate that virus spreading in the vascular wall triggers immune system activation in atherosclerotic plaques and causes endothelial dysfunction. Moreover, productive CMV infection in patients with acute myocardial infarction correlates with the extent of endothelial dysfunction. The mechanisms described by us and other researchers may explain the role of CMV infection in atherosclerosis and development of ischemic heart disease.

Acetylsalicylic Acid Produces Different Effects on the Production of Active Oxygen Species by Activated Platelets in Different Inflammatory Diseases.

We studied the effect of acetylsalicylic acid on ROS generation by platelets in patients after surgical interventions and in patients with bronchial asthma was studied. Platelets stimulated with platelet-activating factor are characterized by weak luminol-enhanced chemiluminescence in healthy people and patients after operations with laparoscopic incisions. Addition of platelet activation factor to platelet samples from patients after open abdominal surgery caused intensive chemiluminescence that was suppressed after platelet incubation with acetylsalicylic acid. At the same time, platelets of patients with aspirin-sensitive asthma did not respond to addition of platelet activating factor, but after incubation with acetylsalicylic acid, an intensive burst of chemiluminescence was detected with a maximum in 5-10 sec after the addition of a platelet-activating factor. In patients with bronchial asthma tolerant to aspirin, platelet activation factor did not induce chemiluminescence irrespective of incubation with acetylsalicylic acid.

Cytomegalovirus in Plasma of Acute Coronary Syndrome Patients.

The relationship between acute coronary syndrome (ACS) and local and systemic inflammation, including accumulation of macrophages in atherosclerotic plaques and upregulation of blood cytokines (e.g., C-reactive protein (CRP)), has been known for more than 100 years. The atherosclerosis-associated inflammatory response has been traditionally considered as an immune system reaction to low-density lipoproteins. At the same time, some data have indicated a potential involvement of cytomegalovirus (CMV) in the activation and progression of atherosclerosis-associated inflammation, leading to ACS. However, these data have been tangential and mainly concerned the relationship between a coronary artery disease (CAD) prognosis and the anti-CMV antibody titer. We assumed that ACS might be associated with CMV reactivation and virus release into the bloodstream. The study's aim was to test this assumption through a comparison of the plasma CMV DNA level in patients with various CAD forms and in healthy subjects. To our knowledge, no similar research has been undertaken yet. A total of 150 subjects (97 CAD patients and 53 healthy subjects) were examined. Real- time polymerase chain reaction (RT-PCR) was used to determine the number of plasma CMV DNA copies. We demonstrated that the number of plasma CMV genome copies in ACS patients was significantly higher than that in healthy subjects (p = 0.01). The CMV genome copy number was correlated with the plasma CRP level (p = 0.002). These findings indicate a potential relationship between CMV activation and atherosclerosis exacerbation that, in turn, leads to the development of unstable angina and acute myocardial infarction. Monitoring of the CMV plasma level in CAD patients may be helpful in the development of new therapeutic approaches to coronary atherosclerosis treatment.

Analysis of Extracellular Vesicles Using Magnetic Nanoparticles in Blood of Patients with Acute Coronary Syndrome.

Extracellular vesicles (EVs) are released from various cell types and play an important role in intercellular interactions. In our study, we investigated abundance of individual EVs in patients with acute forms of ischemic heart disease. Previously, we developed an approach for individual analysis of EVs conjugated with magnetic nanoparticles (MNPs), which was applied in the current study for analyzing phenotypic composition of EVs (by staining for markers CD31, CD41a, and CD63). EVs were isolated using fluorescently labeled MNPs containing anti-CD31, CD41a, or CD63 antibodies and analyzed by combining fluorescently labeled anti-CD41a and CD63, CD31 and CD63, or CD41a and CD31 antibodies, respectively. EVs were analyzed in 30 individuals: 17 healthy volunteers and 13 patients with acute coronary syndrome (ACS). Six and seven ACS patients were with acute myocardial infarction and unstable angina, respectively. It was found that patients with ACS and healthy volunteers contained a dominant subset of EVs expressing surface CD41a antigen, suggesting that they originated from platelets. In addition, the total number of EVs isolated using either of the surface markers examined in our study was higher in patients with ACS compared to healthy volunteers. The subgroup of patients with acute myocardial infarction was found to contain significantly higher number of blood EVs compared to the control group. Moreover, increased number of EVs in patients with ACS is mainly due to the increased number of EVs in the subset of EVs bearing CD41a. By analyzing individual EVs, we found that plasma of patients with ACS, particularly upon developing of myocardial infarction, contained dominant platelet-derived EVs fraction, which may reflect activation of platelets in such patients.

[Immunological Analysis of Human Atherosclerotic Plaques in ex vivo Culture System].

THE AIM OF THE STUDY: to analyze the dynamics of lymphocytic composition of human atherosclerotic plaques in ex vivo culture system. MATERIALS AND METHODS: The study included 15 atherosclerotic plaques obtained from patients who underwent carotid endarterectomy. Plaques were cultured as ring-shaped explants on collagen rafts in culture medium of special composition in CO2 incubator according to the previously developed technique. On day 0, and also on the 4th and 19th days of culture we extracted cells from plaque explants and analyzed B- and T-lymphocytic content of the tissue, as well as the percentage of CD16+ natural killer (NK) cells, using multichromatic flow cytometry. For this purpose we digested the explants with an original enzymatic cocktail, which allows preservation of cell surface markers, and we stained extracted cells with fluorescence-labelled monoclonal antibodies against CD45, CD3, CD19, CD4, CD8, CD16. In addition, we estimated the amount of interleukin 2 (IL-2) and interferon-gamma (IFN-)-producing T-cells by means of flow cytometry. RESULTS: After 4 days of culture the amount of lymphocytes in plaques explants decreased, however live lymphocytes were still preserved (2619.3 [1680.4, 3478.2] cells/100mg tissue). Viable lymphocytes population included T cells (2123.4 [484.9; 3181.2] cells/100 mg tissue), B cells (5.6 [3.4, 27.9] cell/100 mg tissue) and CD16+ NK cells (10.6 [1.8, 23.7] cell/100mg tissue). On the 4th day of culture T cells were presented by CD4+CD8- (797 [475.5, 1000.7] cells/100mg tissue, 37.5 [32.1; 46.3]%) and CD4-CD8+ (686.2 [423.6; 1158.4] cells/100 mg tissue, 45.6 [38.1; 47.9]%) populations. The percentage of CD4+CD8- T cell population decreased compared to the 1st day of culture, and this decrease correlated with the increase in CD4-CD8- T cells content (p<0.05). Additionally, after 4 days of culture we found in tissue explants both CD8+ (17.5[13.3;19.9]%) and CD8- (9.9 [6.4; 14]%) IFN--producing T-cells, however, their percentage, as well as the percentage of IL-2-producing T cells tended to decrease. After 19 days of culture explants of atherosclerotic plaques also contained lymphocytes (2830.1 [2350.3, 5900.2] cells/100mg tissue). Lymphocytes population included T cells (2594.5 [2035.7, 5306.7] cells/100mg tissue), presented by CD4+CD8- (1016.8 [671.2, 2201.7] cells/100mg tissue, 42.3 [34.3; 47.8]%) and CD4-CD8+ (1534.3 [813.8; 2207.2] cells/100mg tissue, 50.8 [45.6; 56.5]%) subsets, B cells (31 [18.3; 64.4] cell/100 mg tissue) and CD16+ NK cells (44.9 [33.4; 138.9] cells/100 mg of tissue). DISCUSSION: An ex vivo model of human atherosclerotic plaque culture that we previously developed enables to preserve viability of various lymphocyte subsets for up to 19 days. We also found that cultured tissue explants retain T cells that can maintain T-helper-1-dependent immune response, which demonstrates inflammation in atherosclerotic plaques. Our results allow to perform experiments on immunological mechanisms of atherogenesis and to develop new approaches for treatment of atherosclerosis, devoted to the suppression of local inflammatory processes in atherosclerotic plaques. CONCLUSIONS: An ex vivo model of human atherosclerotic plaque preserves CD4+CD8- and CD4-CD8+ T cells, B cells, and CD16+ NK cells for a long time. Moreover, after 4 days of culture tissue explants also retain IFN-++ T cells.

[Effect of Remote Ischemic Preconditioning and Postconditioning on Blood Flow in the Infarct-Related Artery and Endothelial Function in Patients With ST-Elevation Myocardial Infarction].

AIM OF STUDY: To assess influence of remote ischemic preconditioning (RIPC) and local ischemic postconditioning and their co-application.

[THE TEST OF GENERATION OF THROMBIN IN DYNAMICS IN PATIENTS AFTER TRANSCUTANEOUS CORONARY INTERVENTION].

The article presents results of observation of generation of thrombin in patients with ischemic heart disease in different terms after transcutaneous coronary intervention. The sampling included 37 patients with stable ischemic heart disease. The control group included 30 healthy individuals. To study system of hemostasis of this category of patients the test of generation of thrombin and its modification with added thrombomodulin were applied for evaluating anti-coagulant activity of system of protein C. The comparison of indicators of test of generation of thrombin in patients with ischemic heart disease before operation and in individuals of control group revealed no reliable differences (p > 0.05). The observation of patients with stable ischemic heart disease in various time-frame after mechanical re-vascularization of myocardium established significant increasing of generation of thrombin and decreasing of anticoagulant activity of system of protein C at 1-3 day after operation (p < 0.05). The positive correlation was established between endogenous thrombin potential and annexin 5, an early marker of dysfunction of endothelium in mentioned time-frame after operation (p = 0.0008; r = 0.57). The significant increasing of content of anti-inflammatory markers of C-reactive protein and fibrinogen was observed at 1-3 day after transcutaneous coronary intervention (p < 0.05). In that way, the study data give evidence to hyper-coagulation in patients with stable ischemic heart disease in early time-frame after operation despite normal values of activated partial thromboplastin time, prothrombin time, D-dimer and applied standard disaggregant therapy.