[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.

[Integral tests of the hemostasis system in assessing the efficiency of acetylsalicylic acid in patients with ischemic heart disease]. / Integral'nye testy sistemy gemostaza v otsenke éffektivnosti atsetilsalitsilovoi kisloty u bol'nykh ishemicheskoi bolezn'iu serdtsa.

Despite the fact that acetylsalicylic acid (ASA) is the "gold" standard for the prevention of cardiovascular complications in patients with coronary heart disease (CAD), a number of patients still have risks of atherothrombosis. In the present study, the antithrombotic effect of ASA in patients with CAD was assessed in platelet-rich plasma (PRP) using integral tests of the hemostasis study: the T-TAS system (Total Thrombus-formation Analysis System) and the thrombin generation test (TGT). The study involved 34 patients with stable CAD (11 women, 23 men) and people (15 women, 18 men) in the control group. As a result of assessing the activity of thrombus formation using the T-TAS system, a significant decrease in the area under the curve (AUC10) was found in the group with CAD patients compared with the control (135.6 [88.0-222.3] and 260.5 [217.3-301.9], respectively, p.

[The possibility of using neuron-specific enolase as a biomarker in the acute period of stroke]. / Vozmozhnost' ispol'zovaniia neiron-spetsificheskoi enolazy kak biomarkera v ostrom periode insul'ta.

The article presents a review of the literature on neuron-specific enolase (NSE) as a biomarker of stroke. It is shown that NSE does not allow differentiation of the ischemic and hemorrhagic process in stroke, but is suitable for determining the extent of brain tissue destruction both in the first hours of stroke and in the dynamics. The HSE analysis can be useful for monitoring the course of the disease, control of the dynamics of the pathological process, including when the size of the lesion increases, for evaluating the effectiveness of therapy and as a prognostic biomarker.

[Anticoagulant activity of direct factor Xa inhibitors as a tool to ensure the effectiveness and safety of drugs intake].

The thematic review presents modern solutions using oral anticoagulants with a focus on direct coagulation factor X inhibitors. It contains information about the pharmacodynamics and pharmacokinetics of apixaban and rivaroxaban against the background of different drug intake regimens - twice and once per day. There are shown studies of concentration dynamics and the corresponding functional response, measured using the integral method - the thrombin generation test, which is widely used in scientific research to describe hemostatic processes based on an objective quantitative assessment of the thrombin formation - a key coagulation cascade serine protease. The logical relationship between the pharmacodynamics of anticoagulant action and the clinical presentation of the effectiveness and safety of drugs is traced. The review provides links to actual literature and current clinical guidelines.

[Increase in GP IIb-IIIa and P2Y12 Receptors in Activated Platelets as the Possible Indicator of de novo Protein Synthesis].

Although platelets lack nuclei, they are capable of de novo protein synthesis. We speculate that key platelet receptors are involved in the regulation of this process, and the changes in their number indicate the de novo protein synthesis in platelets. The object of our study was native platelets obtained from healthy donors. Using flow cytometry and Western blot, we determined the number of GP IIb-IIIa receptors (fibrinogen receptor) and P2Y12 receptors (ADP receptor) on the surface of platelets upon their activation with ADP and collagen. To verify the approaches and techniques used, we studied IL-1ß protein, which was previously shown to be synthesized de novo in activated platelets. GP IIb-IIIa receptor numbers correlate with the number of P2Y12 receptors on the cell surface (R = 0.45, p = 0.03). It was demonstrated that the platelet receptor numbers are higher on the surface of the cells with high functional activity. According to the data obtained by Western blot, upon the cell activation with ADP, the number of GP IIb-IIIa and P2Y12 receptors increases, which may serve as evidence of these proteins being synthesized in the activated platelets. It was observed that the level of P2Y12 and IL-1ß was lower in the samples where GP IIb-IIIa receptor was blocked by the selective inhibitor, i.e., the Fab fragment of the antibodies that specifically recognizes the GP IIb-IIIa complex. This suggests the important role of GP IIb-IIIa receptor in the regulation of protein synthesis.

[The molecular mechanisms of platelets activation in patients with cerebrovascular disease].

Cerebrovascular disease is a main cause of mortality and one of the big medical problems. After the vascular wall's damage the endothelial cells secrete the von Willebrand factor which then connects with its platelet's receptor GP Ib-V-IX. There are two polymorphisms Thr145Met and T(-5)C of the GP Iba gene associated with arterial thrombosis development. Also the difference in platelets' genes expressions was shown in patients with various clinical course of ischemic heart disease. The aim of this study was to investigate the role of platelet's receptor for von Willebrand factor in platelets' activation in patients with cerebrovascular disease. 123 patients with cerebrovascular disease and 97 healthy donors were included into the study. We analyzed the level of receptor for von Willebrand factor on platelet's membrane by flow cytometry, Thr145Met and T(-5)C GP Iba polymorphiams by PCR-RFLP, the GP Iba gene expression by RT-PCR and ADP-induced platelet aggregation by Born method. We have shown: 1) the 145Met GP Iba allele prevalence in patients with atherotrombotic stroke development due to macroangiopathy; 2) the pre-mRNA transform into the mature mRNA in activated platelets and this process may be stopped by the antiplatelet therapy by acetylsalicylic acid.

[The clinical laboratory diagnostic as a part of clinical medicine.]

The rapid development of laboratory technologies and their translation directionality permit developing principles of personified medicine. The article considers new paradigm of manpower policy for developing necessary level of clinical laboratory consultation.

[Laboratory assessment of haemostatic parameters in patients taking a direct thrombin inhibitor].

The problem of prevention and treatment of thromboembolic complications has a significant place in clinical practice for many years. The gold-standard agents in long-term protection from embologenic strokes, secondary prevention of venous thromboses and embolisms still remain vitamin K antagonists (in Russia - warfarin). However, despite high efficacy, administration of warfarin is fraught with dangers and associated with a series of inconveniences. A direct thrombin inhibitor, dabigatran etexilate (hereinafter referred to as dabigatran) was approved in the Russian Federation for prevention of thromboembolic complications in orthopaedic practice (2009), for prevention of ischaemic embologenic stroke in atrial fibrillation (2011) and for treatment of recurrent thrombosis of deep veins and pulmonary artery thromboembolism (2014). A characteristic feature of a therapeutic agent possessing an anticoagulation effect is correlation between intensity of hypocoagulation and haemorrhage. The effect of dabigatran on the laboratory parameters of haemostasis has been studied insufficiently, with no practical guidelines on assessing these alterations for prediction of the risk for haemorrhagic and thromboembolic complications. The present study included a total of 65 patients with non-valvular aetiology atrial fibrillation, taking dabigatran during from 6 to 18 months. All patients underwent laboratory assessment of the coagulation level and measuring blood coagulation activation markers in dynamics 10-14 days, 1, 6, 12 and 18 months after taking the agent. Thromboembolic and haemorrhagic risks were also assessed. It was revealed that administration of dabigatran leads to alterations in the main parameters of coagulogram. Determination of prothrombin (in % according to Quick's method) and activated partial thromboplastin time may be used for qualitative assessment of hypocoagulation. During the follow up period no statistically significant changes in the coagulation activation markers level were observed.

[Gender differences in clinical and hemoreologic disorders in elderly patients with non-cardioembolic ischemic stroke].

Clinical manifestation and platelet homeostasis parameter results in elderly patients with atherothrombotic and lacunar strokes are presented. There is platelet activation in these patients manifested by increasing of P-selectin expression in response to adenosindiphosphate induction on flow-cytometry. Agregometry method was not informative for platelet activity estimation in these patients. Von Willebrand factor receptor expression on the platelet may have prognostic value for acute period. The fibrinogen receptor expression on the platelet was increased in women with stroke comparing with men.

[Apoptosis of peripheral blood lymphocytes in patients with LRRK2-assoctated Parkinson's disease].

Mutations in the Leucine Reach Repeat Kinase 2 (LRRK2) gene are the most frequent cause of familial Parkinson's disease (PD). Although the precise physiological and pathological role of LRRK2 is unclear, a direct link between mutant LRRK2 and apoptosis has been suggested. Using flow cytometric analysis (PI+Annexin V(FITC)) we showed increased spontaneous apoptosis of peripheral blood lymphocytes in patients with LRRK.2-associated PD compared to controls after 24 (P < 0.016) and 48 (P < 0.031 ) h of incubation (5 % CO2, 37 degrees C). We found the increased FAS mRNA level in peripheral blood lymphocytes of patients with LRRK2-associated PD compared to controls (P < 0.05) and to sporadic PD (sPD) (P < 0.002). Significant difference in FAS expression between patients with LRRK2-associated PD and controls remained after three years and was detected after 1 and 24 h during lymphocyte incubation (P < 0.03 and 0.05, respectively). Increased spontaneous lymphocytes apoptosis along to increased FAS expression in patients with LRRK2-associated PD suggest that LRRK2 mutations may lead to the activation of extrinsic apoptotic way.

[The preanalytical stage of studying the hemostatic system].

The efficiency of applying precision devices and multiple laboratory methods devised to reveal the causes of bleeding and thromboses significantly depends on whether a wide range of rules of the preanalytical stage of the study is observed. The paper gives detailed information on conditions for taking blood samples, their transport, and preparation for examination of the hemostatic system. The given information is based on the worldwide recommendations and many years' experience gained by the authors at the laboratory of hemostasis.

[Clinical and laboratory criteria for diagnosis in antiphospholipid syndrome: what is it important for a practitioner to know (a lecture)].

The antiphospholipid syndrome (APS) is antibody-induced thrombosis, whose diagnostic basis is the obligatory presence of serological markers along with clinical manifestations. Its laboratory markers are antiphospholipid antibodies (aPAs), the highest informative value has been proven for lupus anticoagulant (LA), cardiolipin antibodies, fl-glycoprotein 1. LA should be determined at a laboratory in accordance with the recommendations of the International Society on Thrombosis and Hemostasis. Obstetric abnormality in APS should be regarded as a thrombotic complication and its management in this case does not differ from that and prevention of thrombosis, aPAs may be detectable in various illnesses, but their blood presence is not an indication for immunosuppressive therapy. The paper describes a case of primary APS in a female patient.

[Genetic variants of platelet ADP receptor P2Y12 associated with changed platelet functional activity and development of cardiovascular diseases].

The key role in platelet aggregation is played by the platelet ADP receptor P2Y12, which is the target for antiaggregant drugs, clopidogrel and ticlopidine. At present, only sporadic data on genetic variants of platelet ADP receptor P2Y12 are available from literature, and their association with thromboembolic and cardiovascular diseases still remains obscure. Analysis of the group of subjects with high platelet reactivity resulted in identification of two nucleotide substitutions, C18T and G36T, in the coding region of the P2Y12 gene. The frequency of the P2Y12 T1 8 allele was higher in control group than in the group of patients survived from myocardial infarction at the age under 45 years (39% versus 28%, respectively, P = 0.04). Moreover, in the T18 carriers, platelet aggregation activity was lower than in the carriers of the wild-type genotype (0.84 +/- 0.05% versus 1.01 +/- 0.08%, respectively, P = 0.03). In the group of patients with early myocardial infarctions, a tendency towards the increased frequency of 16T allele in comparison with control group (20 and 12%, respectively, P = 0.07) was observed. The rate of ADP-induced platelet aggregation in the carriers of 16T allele from the control group was somewhat higher than in the subjects with the GG36 genotype (1.31 +/- 0.16% versus 1.12 +/- 0.06%, respectively, P = 0.07). The nucleotide substitutions identified were in absolute disequilibrium, i.e., allele T18 conformed to allele G36. On the contrary, allele C18 conformed to allele T36. Haplotype T18G36 was found to be responsible for the decreased risk of myocardial infarction and decreased platelet reactivity. It is suggested that polymorphisms of the P2Y12 gene identified can be used for determination of the risk group for myocardial infarction in the young males.

[Evaluation of D-dimer in outpatients: clinical value and specific features].

D-dimer as an activation marker of coagulation and fibrinolysis is a recognized diagnostic criterion of deep vein thrombosis, pulmonary thromboembolism, and disseminated intravascular coagulation. In recent years, this laboratory test has been most frequently used for other purposes: to detect the activation of coagulation, to predict the course of diseases, and to determine the duration of anticoagulant therapy. Our investigation examined 1514 D-dimer measurements in 1370 outpatients without acute abnormalities, including 72 patients receiving warfarin and 32 patients after myocardial revascularization. 36.1% of cases were found to have values of more than 0.5 mkg/ml. Adequate anticoagulant therapy (INR 2-3) caused a reduction in the level of D-dimer that is an important additional laboratory test for the evaluation of antithrombotic defense. Further investigations are needed to determine cutoff values for various clinical situations.

Participation of IIb-IIIa glycoprotein in spontaneous platelet aggregation.

In some patients with stable and unstable angina pectoris and in some donors without clinical manifestations of cardiovascular diseases and other pathologies, spontaneous platelet aggregation was completely suppressed by glycoprotein IIb-IIIa antagonists blocking the interaction of this glycoprotein with fibrinogen. Antibodies inhibiting binding of glycoprotein Ib with von Willebrand factor had no effect on the level and rate of spontaneous platelet aggregation. In the donor group, the level of spontaneous aggregation was almost 1.5-fold higher in persons with a certain genetic polymorphism (Leu-->Pro substitution in position 33 of glycoprotein IIIa). The level of spontaneous aggregation correlated with the amount of glycoprotein IIb-IIIa on the platelet surface (r = 0.41).

[Novel mutation of the GPIIIa gene in the Russian population, Leu40Arg, linked to the Leu33Pro].

Glycoprotein IIb/IIIa complex, a platelet surface fibrinogen receptor, plays a key role in producing primary hemostasis. At present, only a single mutation in the GPIIla gene, Leu33Pro, and a single mutation in the GPIIb gene, lle843Ser, has been described. The mutations are known to enhance signaling functions of the receptor and are associated with the development of arterial thromboses. In the present study, we describe a novel GPIIIa mutation, which is T to G nucleotide substitution in position 1585, resulting in the replacement of Leu for Arg in position 40 of the amino acid sequence of the protein.

[Clinical value of allele variants of cytochrome CYP2C9 gene for anticoagulation therapy with warfarin].

Warfarin is metabolized by cytochrome CYP2C9 and its pharmacokinetic properties depend on structural polymorphisms of CYP2C9 gene. We studied frequencies of allele variants of CYP2C9 gene and associations of individual reaction to warfarin intake with genotype of CYP2C9 gene. Population frequencies of CYP2C9x1, CYP2C9x2, CYP2C9x3 alleles of CYP2C9 gene in St-Petersburg were 82.66, 11.11, and 6.32%, respectively. Carriers of CYP2C9x2 and CYP2C9x3 alleles more rapidly achieved therapeutic levels of hypocoagulation and required significantly lower weekly doses of warfarin.

[Models of inter-group difference of the homeostasis system in patients with artificial heart valves and the set of laboratory parameters]. / Modeli mazhgruppovykh razlichii sostoianiia sitemy gemostaza u bol'nykh s isskustvennymi klapanami serdtsa po kopleksu laboratornykh pokazatelei.

Patients with artificial heart valves (AHV) belong to a unique multi-component model of the homeostasis system presenting an interaction of activated coagulation processes and pharmacological effects, on the one hand, and genetic peculiarities functioning homeostatic mechanisms, on the other hand. The described method was evaluated on the basis of multi-factor mathematic analysis by information-metry. The data obtained provided for isolating the most information-dense laboratory indices and some genetic polymorphisms describing the shaping of continuous intravascular coagulation of stage III in AHV patients. The clinicians and laboratory experts are offered a practical instrument for the diagnosis (prognostication) of the above condition. It was proven as necessary to examine (by the morphofunctional method) the intravascular activation of platelets and to determine the D-dimer in the remote period after prosthesis of heart valves.