[A clinical case of reverse left ventricular remodeling in patient with pathogenic TTN mutation. Case report].

Dilated cardiomyopathy (DCM) is a leading cause of heart failure, sudden cardiac death, and heart transplantation in young patients. The causes of DCM are varied and include genetic factors and metabolic, infectious, toxic and others factors. Today it is known that germline mutations in more than 98 genes can be associated with the occurrence of DCM. However, the penetrance of these genes often depends on a combination of factors, including modifiable ones, i.e. those that change under the influence of the environment. About 20-25% of genetically determined forms of DCM are due to mutations in the titin gene (TTN). Titin is the largest protein in the body, which is an important component of the sarcomer. Although titin is the largest protein in the human body, its role in the physiology of heart and disease is not yet fully understood. However, a mutation in the TTN gene may later represent a potential therapeutic target for genetic and acquired cardiomyopathy. Thus, the analysis of clinical cases of cardiomyopathy in patients with identified mutations in the TTN gene is of great scientific interest. The article presents a clinical case of manifestation of DCM in patient with a revealed pathogenic variant of mutation in the gene TTN and reverse left ventricular remodeling of the against the background of optimal therapy of heart failure in a subsequent outpatient observation.

[Actual nutrition in adults with familial hypercholesterolemia].

Familial hypercholesterolemia (FH) is a highly atherogenic, genetically based lipid disorder. For patients with FH, dietary modification is the cornerstone of complex lipidlowering therapy. The aim of the research was to assess the actual nutrition in adults with familial hypercholesterolemia. Material and methods. The study included 100 patients over 18 years old (including 46% men) with "probable" or "definite" FH according to the Dutch Lipid Clinic Network or Simon Broome criteria from the GENMOTIV-FH study (ClinicalTrials: NCT04656028) in 2019-2021. Actual nutrition was assessed using the 24-hour dietary recall method. The frequency of the main meal groups' consumption and food-related behavior were assessed using a questionnaire method. The data are presented as the median [Q25; Q75]. Results. The study showed the excess consumption of protein (19.3 [16.7; 24.0] in men and 18.6% [13.6; 24.3] in women, p=0.592), total fat (35.1 [29.4; 41.0] in men vs 39.2% [33.2; 47.5] in women, p=0.018), including saturated fatty acids (9.6 [4.7; 13.0] vs 10.4% [7.5; 14.2], respectively, p=0.151), and cholesterol (265.8 [188.8; 521.9] mg/day in men vs 282.1 [147.2; 542.8] mg/day in women, p=0.936). Consumption of total carbohydrates (44.3 [37.2; 50.0] vs 39.6% [30.1; 48.8], respectively, p=0.100) and fiber (10.7 [7.3; 13.3] g/day in men vs 11.5 [7.9; 13.9] g/day in women, p=0.372) was insufficient. Only 47.9% of patients consumed vegetables daily, 39.1% - fruits and berries. The majority (64.5%) of patients with FH preferred high-fat cheese (>=25%). Cottage cheese of >=5% fat content preferred 52.7% of patients. The daily poultry consumption was more than red meat (19.3 vs 4.3% respectively, p=0.003). Regularly included fish in their meal 53.8% of patients. Conclusion. The actual nutrition in adults with FH does not match international guidelines. The results highlight the importance of dietary interventions for patients with FH.

[Familial left ventricular noncompaction: phenotypes and clinical course. Results of the multicenter registry].

AIM: To analyze and demonstrate various phenotypes in patients with familial left ventricular noncompaction (LVNC). MATERIALS AND METHODS: In 2013 was created a multicenter registry of LVNC patients. On its basis 30 families with a familial LVNC were selected. RESULTS: 30 LVNC families were selected from the register. From a total of 115 people (probands and relatives) in 71 (61.7%) LVNC was diagnosed (30 probands and 41 relatives with non-compact myocardial criteria). The most common type of remodeling in patients was the dilated type (DT) (n=30), the isolated LVNC with preserved ejection fraction (EF) was slightly less common (n=23), and the hypertrophic type (GT) was detected in 8 patients. 4 patients were diagnosed with the isolated LVNC with a reduced EF. 3 patients were with a combination of non-compact myocardium with congenital heart disease and with a combination of DT and GT (DT+GT). During the analysis of cases a combination of different phenotypes in the same family was observed. The largest number of families was diagnosed with a combination of DT and the isolated LVNC with preserved EF. The development of cardiovascular complications was associated with DT. CONCLUSION: Family cases of LVNC had different types of myocardial remodeling and variants of clinical course. In one family a combination of different types of left ventricular remodeling is possible. DT is associated with the most severe clinical manifestations. The clinical picture of the isolated LVNC with preserved EF, is the most favorable, but in rare cases, serious clinical manifestations were observed.

Actual approaches to the transportation of biological samples at low temperatures.

Taking into account the impact of shipment method of biosamples is necessary for obtaining high-quality biological samples in biobanking and laboratory research. The impact of liquid nitrogen, dry ice and cold accumulators on the quality of biological markers was considered, as well as recommendations to reduce the impact of these methods of shipment. The liquid nitrogen provides the best preservation of samples, however, dry ice is used much more often during their transportation. When transporting certain types of cells using dry ice, there is the way to use CryoStor CS1 and Cell Banker 1 cryoprotectors. The dry ice has a significant effect on both the pH of liquid biological samples and the coagulological parameters of plasma samples. The penetration of CO2 into the sample leads to changes in the parameters of PTT and APPT, as well as to decrease the protein C and fibrinogen level under certain conditions. Serum and plasma samples exposed to dry ice for more than 16 hours should be thawed open at room temperature, or instead of it should be kept at -80 °C for 24 hours to avoid changes in coagulation parameters, The use of cold accumulators is unacceptable for long-term shipment of serum and plasma containing unstable biomarkers because of insufficiently low temperature (increase over time to -25 °C and above). Besides, metal pellets can be used as cold storage batteries at low temperatures (up to -80 ° C), but they are not as effective as dry ice, since it is able to hold the required temperature for much longer.

ISO 20387 biobanking standard. Analysis of requirements and experience of implementation.

Currently one of the most important problems facing biobanking specialists is the standardization of biobanks operation. Close attention is paid to this issue by international biobanking organizations, such as ISBER and BBMRI-ERIC, which develop regulatory documentation in this area. The article provides examples of standardization tools - implementation of the ISO 9001 quality standard and ISBER Best Practices. General information about the development, scope, and structure of the ISO 20387 standard is provided. The standard does not provide ready-made solutions and does not contain specific requirements for storage temperature or biosamples processing in biobanks, allowing each biobank to adapt its own management system to existing conditions and needs. The standard contains requirements for both the organization of the biobanking and the supporting processes - personnel competence; requirements for biological safety; infrastructure management, including equipment used by the biobank, environmental parameters that affect the storage of biomaterial. The standard contains requirements for the quality management system of biobank, as a necessary element of the organization of any biorepository. At the initiative of the Russian National Association of biobanks and biobanking specialists (NASBIO), development of the Russian standard GOST R ISO 20387 «Biotechnology. Collection and storage of biological samples in biobanks. General requirements¼ is included in the plan of the National Standardization Program for 2020 by order of Rosstandart No. 2612 of 11/01/2019. Implementing quality standards is a long and painstaking process that requires the involvement of all employees and certain resources. However, the effectiveness of strict compliance exceeds the cost of developing, implementing and maintaining management systems, as it significantly increases the confidence of researchers in the work of biobanks, guarantees high quality of biospecimens and associated data, and creates opportunities for cooperation, both at the national and international level, based on the application of common quality standards in the work.

[Quality management in biobank. World tendencies and experience of biobank of FSI «NMRC for preventive medicine¼ of the Ministry of healthcare of Russia.]

This paper provides several definitions of the term "biobank"; a list of standards developed by the International Organization for Standardization (ISO) applicable to the activities of biobanks; analyzes the legal and ethical requirements; the Russian legal framework in the field of biobanking, the best international practices and recommendations; describes the experience of the development and implementation of quality management systems according to ISO 9001 in Biobanks established in different countries, and the experience of the Bank of Biological Material of the National Medical Research Center for Preventive Medicine of the Ministry of Healthcare of Russia. The ISO 20387 Biobanking Standard, released in August 2018, combined the knowledge and experience of specialists from around the world and defined the general requirements that must be fulfilled by biobanks and repositories wishing to guarantee their customers the high quality of the preanalytical stage of scientific research, biological samples and associated data. In 2019, the Russian version of this standard is expected. Documents of the quality management system provide the reproducibility of activities on the main storage processes and facilitates the process of incorporating a new employee; conducting internal and external audits; Biobank knowledge management - continuous staff education. The introduction of an effective quality management system into biobank activity warrants the high quality of biological samples, the standardized pre-analytical stage, reliable, regulated long-term storage of biomaterial and related information for use in research purposes today and in future.

[Current best practices and biobanking recomedations.]

The biobank is a structure established with the goal of long-term responsible storage of biological samples and the associated data for their further use in scientific and clinical research. The objectives of biobanking are the creation of unified recommendations on: the planning of premises and the selection of equipment for storage; development of management methods and staff training; standardization of methods for the collection, shipping, processing and storage of biomaterial of various origins, as well as methods for quality control and validation of the applied methods; creation and use of databases of information accompanying biospecimens. The lack of common standards for conducting the preanalytical phase has been the cause of low accuracy and poor reproducibility of research results. To date, a large number of guidelines and best practices have been published that provide an answer to a wide range of problems in organizing the biobanking process. The article provides an overview of the most famous biobanking guidelines that can be used to solve various research problems. Biobanking in Russia is actively developing. Since 1996 there is a work on the legislative regulation of biobanking activities, as a result of which a number of regulatory documents have been issued. An important stage in the development of biobanking in Russia was the establishment of the "National Association of Biobanks and Biobanking Specialists" (NASBio) in 2018, which included representatives of medical and research institutions, commercial firms, and qualified specialists in the field of biobanking. One of the key tasks of NASBio is the adaptation and implementation of the best biobanking practices in Russian research institutes and centers. The use of modern guidelines and best practices on biobanking will lead to an increase in the quality of research and publications.

Association between polymorphisms of eNOS and GPx-1 genes, activity of free-radical processes and in-stent restenosis.

Our aim was to examine correlations between polymorphisms in five antioxidant enzymes genes, activity of free-radical processes, and the risk of restenosis after coronary artery stenting with bare metal stents (BMS). A total of 101 male patients who underwent intracoronary stenting using BMS and coronary angiography follow-up of 6 months were enrolled in: group with in-stent restenosis (n = 44) and without restenosis (n = 57). The content of lipoperoxides and malondialdehyde (MDA) in Low-density lipoprotein (LDL), activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) in erythrocytes, and genotypes polymorphisms of the CAT gene (-262C/T), paraoxonase-1 (PON-1) gene (163T/A and 575A/G), endothelial nitric oxide synthase (eNOS) gene (298G/T (rs#1799983) and -786T/C), GPx-1 gene (599C/T (rs#1050450)), and glutathione-S-transferase (GSTP) gene (313A/G) were determined. In carriers of the minor allele of 599C/T polymorphism of the GPx-1 gene, activity of GPx in erythrocytes was lower by 17 % than in wild allele homozygotes, while the content of lipoperoxides in LDL was higher by 74 %. T-allele of 599C/T polymorphism of the GPx-1 gene (OR = 2.9; 95 % CI: 1.23-6.82) and T-allele of 298G/T polymorphism of the eNOS gene (OR = 2.79; 95 % CI: 1.17-6.66) were associated with the risk of in-stent restenosis. Minor alleles of polymorphisms 298G/T of the eNOS gene and 599C/T of the GPx-1 gene are associated with an increased risk of in-stent restenosis. Minor allele of the GPx-1 gene 599C/T polymorphism leads to a decrease of the GPx activity and increase of the activity of free-radical processes.

Pro198Leu polymorphism of GPx-1 gene and activity of erythrocytic glutathione peroxidase and lipid peroxidation products.

We studied the relationship between the GPx-1 gene Pro198Leu polymorphism genotype and activity of free-radical processes. In patients with documented CHD, the content of lipoperoxides and MDA in LDL, activity of glutathione peroxidase in erythrocytes, and genotype of GPx-1 gene Pro198Leu polymorphism were determined. In carriers of the minor allele, activity of glutathione peroxidase in erythrocytes was lower by 17% than in wild allele homozygotes, while the content of lipoperoxides and MDA in LDL was higher by 74 and 27%, respectively. We concluded than free-radical oxidation processes are more pronounced in carriers of the minor allele of GPx-1 gene Pro198Leu polymorphism.

[Effect of polymorphisms of cytochrome P450 and apolipoprotein E genes on therapeutic efficacy of statins].

Statins are widely used in clinical practice for lowering of levels of atherogenic blood plasma lipids and treatment of atherosclerosis. Variability of response of the body to these drugs might be determined by genetic factors (gene polymorphisms) related to metabolism of drugs. Among them central place belongs to enzymes of subfamily 3A of cytochrome P450 (CYP). In this review we present results of studies assessing effect of various allele variants of CYP3A4 and CYP3A5 on efficacy and tolerability of atorvastatin, lovastatin,, and simvastatin in different populations of patients. We also present data on populational frequency of genetic polymorphisms under study. In addition we cover the problem of possible influence of apoE genotype on efficacy of statins. The available data do not allow yet to recommend pharmacogenetic testing for wide clinical practice.

[Diagnostics of moderate and high hypertriglyceridemia in patients in polyclinic practice: primary and secondary lipid metabolic disorders].

AIM: To define clinical and biochemical differences in groups of patients with moderate (< or =4.5 mmol/l and high (more than 4.5 mmol/l) blood triglyceride (TG) levels. To define the markers of biochemical and lipid parameters that could specify an algorithm for the differential diagnosis and treatment of different forms of hypertriglyceridemia. SUBJECTS AND METHODS: Patients (96 (54%) females) aged 12 to 71 years (median 50 years; quartiles, 41-61 years) with a TG level of more than 23 mmol/l and the following diseases: coronary heart disease (CHD) (44.8%), myocardial infarction (13.5%), arterial hypertension (87.9%), xanthomas (36.5%), and a family history of diseases (51%). The diagnosis of hyperlipidemia included a classical algorithm for clinical, biochemical, and clinicogenealogical examinations. Extended biochemical blood analysis, the determination of lipoprotein cholesterol (C), TG, low-density lipoprotein C, lipid electrophoresis, and assay of apolipoproteins A1, B-100, E, and C3 were made. RESULTS: The groups with moderate (< or =4.5 mmol/l and high (more than 4.5 mmol/l) blood triglycerides showed no differences in age and gender, systolic and diastolic blood pressures, the incidence of coronary heart disease, arterial hypertension, peripheral artery atherosclerosis, cardiac arrhythmias, and xanthomas. There was a significant correlation of high TG levels with smoking (a risk factor) and with the indicators of other metabolic disturbances--total C, chylomicrones, lipoprotein(a), LP-E-total, LP B:E, LP-C3-total, and LP-C3, which determined the impact of nutrition (and the development of pancreatitis), but also had a hereditary predisposition through the polygenic mechanisms of gene expression under the influence of a number of factors. CONCLUSION: Higher TG levels correlated with the parameters, the diagnosis of which makes it possible to reveal additional metabolic disturbances via environmental and polygenic mechanisms.

[Disorders of lipid metabolism in patients with cholelithiasis and gallbladder cholesterosis depending on apolipoprotein E genotype].

The work was aimed to study blood lipid spectrum in 133 patients with cholelithiasis (CL) and 159 with gallbladder cholesterosis (GC) as well as apoE genotypes (based on restriction fragment polymorphism) in 49 and 36 respectively. Lipid composition was shown to significantly differ in the two conditions. LDL cholesterol was increased in GC and TG in CL. A rise in LDL cholesterol in both groups was apparent before the age of 30 yr (34.6 +/- 8.4 and 52.6 +/- 12.9% respectively), that in TG and VLDL after 40 yr. E3/3 genotype (norm) was identified in 75.5 +/- 6.2% of the patients with CL and in 83.4 +/- 6.2% in those with GC (p < 0.05). e4 allele (mutation) equally frequently occurred in 10.2 +/- 4.3 and 8.1 +/- 4.5% of patients with CL and GC (p > 0.5), e2 allele in 14.5 +/- 5.0 and 8.1 +/- 4.5% (p < 0.05). These data suggest that patients of both groups equally frequently suffered disturbances in metabolism of saturated (e2 allele) and polyunsaturated (e4 allele) fatty acids predisposing for hypercholesterolemia and hyperlipidemia. They explain why CL is frequently associated with cholesterosis and GC with the formation of caliculi. However, the absence of significant correlation between CL, GC and alleles e2, e4 suggests participation of other factors in pathogenesis of these diseases (LP(a), LDL heterogeneity).

Binding of pyridoxal, pyridoxal 5'-phosphate and derived hydrazones to bovine serum albumin in aqueous solution.

In the present paper, the kinetics of a reaction between bovine serum albumin (BSA) and pyridoxal, pyridoxal 5'-phosphate was studied, apparent rate constant of product formation and dissociation as well as binding constants were determined. Pyridoxal 5'-phosphate hydrazones of isonicotinic, picolinic, 2-furoic, thiophene-2-carboxylic, pyrazinoic acids binding to BSA was studied by spectrofluorimetry, stability constants of the associates were calculated from experimental data using maximal likelihood approach. The changes in the secondary structure of BSA induced by hydrazones addition were studied by IR spectroscopy. New freely available software for curve fitting was developed as a part of the software kit designed for the solution chemistry and used for a specific problem of this study, IR spectra processing.

[Familial hypercholesterolemia in Russia: genetic and phenotypic characteristics].

AIM: To make qualitative and quantitative analyses of phenotypical characteristics and to study a spectrum and frequency of mutations in LDLR and APOB genes in patients with familial heterozygous hypercholesterolemia (FHHC). MATERIAL AND METHODS: Clinical symptoms of FHHC were studied in males and females. Mutations were detected with PCR, analysis of SSCP of all the exones of LDLR gene and a fragment of exone 26 of APOB gene with subsequent sequestration of DNA fragments with anomalous electrophoretic motility, analysis of restriction fragments length polymorphism. RESULTS: LDLR gene mutations were detected in 50%, of APOB gene in 2.6% patients with FHHC, 70% of LDLR gene mutations have never been discovered before. Three known mutations were detected in the APOB gene: R3500Q (1.9% cases), H3543Y (0.55%), R3531C (0.15%). Incidence of coronary heart disease in untreated FHHC patients is 61.5%, of myocardial infarction--31%. Life span of both males and females with FHHC was subnormal, especially of men (median: 53 years in 95% CI, 49.2-56.8 years and 62 years in 95% CI 59.2-64.8 years, respectively). Incidence rate of basic clinical symptoms increased with age and significantly correlated with LDLP cholesterol. CONCLUSION: Frequency and severity of clinical symptoms and complications in FHHC and in Russian population agree with those of the European countries. The same occurs with frequency and mutations of the APOB gene, while mutations of the LDLR gene in 70% cases are unique for Russian population and are not described in other countries. This makes impossible to use foreign test kits for FHHC diagnosis in Russia.

KEV: A free software for calculating the equilibrium composition and determining the equilibrium constants using UV-Vis and potentiometric data.

Present paper reports on an algorithm for calculating the equilibrium composition of several compounds mixture if their total (equilibrium) concentrations and the equilibrium constants of reactions between them are known. The algorithm for determining the unknown equilibrium constants from UV-Vis or potentiometric experimental data using minimizing function (maximal likelihood method) is described. The recommendations on the evaluating of equilibrium constants from experimental results are given. The algorithms described are realized as free-of-charge distributed software/scripts bundle/source code.

[Phenotypical features of a heterozygous form of familial hypercholesterolemia].

AIM: To conduct a quantitative and qualitative analysis of phenotypical manifestations in patients with a heterozygous form of familial hypercholesterolemia (FHC) and to reveal factors involved in their development. MATERIAL AND METHODS: A total of 247 patients with a clinical diagnosis of heterozygous FHC participated in the trial. Clinical manifestations of the disease in men and women were analysed and compared. Blood lipids were compared to those in the controls. A correlation analysis was used to reveal correlations between symptoms of the disease and lipid levels in the blood. RESULTS: Tendon xanthomas were most frequent (79%) clinical sign with location primarily in Achilles' tendon. Incidence of basic clinical manifestations increased with age and significantly correlated with LDLP cholesterol. Two clinical signs were seen in 1/3 of the patients, three--in 13% (sex differences were insignificant). Mean levels of total cholesterol and LDLP serum cholesterol in heterozygous patients were 1.9 and 2.5 times higher than in the controls. Total cholesterol was significantly higher in women. A mean level of HDLP cholesterol was significantly lower while triglycerides were higher than in the control group. The disease symptoms manifested in men 5 years earlier than in women, FHC was diagnosed in men 7.5 years earlier. CONCLUSION: Patients with heterozygous FHC are characterized by higher levels of LDLP cholesterol, lower level of HDLP cholesterol and higher triglycerides in the serum than in healthy controls. Sex-related differences by severity and prevalence of basic symptoms in heterozygous FHC patients were not found. The time of clinical symptoms appearance and diagnosis evidences for more rapid progression of the disease in men.

[Genetic factors of risk of ischemic heart disease development in patients with familial hypercholesterolemia].

UNLABELLED: Ischemic heart disease (IHD) develops in patients with familial hypercholesterolemia (FHC) 15-20 years earlier than in general population. However age of onset of the disease, its clinical manifestations are variable and not completely determined by cholesterol level and class of low density lipoprotein receptor mutations. AIM: To elucidate associations of some auxiliary genetic factors -- such as C151565T, C677T, R353Q polymorphisms of glycoprotein IIIa (GPIIIa), methylenetetrahydrofolate reductase (MTHFR) and coagulation factor VII genes, respectively, -- with the presence of IHD in patients with FHC. MATERIAL: Patients with clinical diagnosis of heterozygous FHC (n=198) with (n=106) and without (n=92) IHD. RESULTS: Patients with compared with those without IHD had similar frequency of T-allele of MTHFR gene (p=0.519), more often had T-allele of GPIIIa gene (23 and 12.5%, respectively, p=0.009), and less often -- Q-allele of factor VII gene (13 and 21%, respectively, p=0.048). Multifactorial analysis showed that risk of IHD was higher in patients with TT compared with CC genotype of the GPIIIa gene (OR 1.53, 95%CI 1.12-2.3), and lower in patients with RQ and QQ compared with RR genotype of factor VII gene (OR 0.41, 95%CI 0.19-0.75). CONCLUSION: In patients with FHC polymorphisms in factor VII and GPIIIa genes but not C677T polymorphism of MTHFR gene were associated with the presence of IHD.

[Low density lipoprotein receptor gene mutations in patients with clinical diagnosis of familial hypercholesterolemia.]. / Mutatsii gena retseptora lipoproteinov nizkoi plotnosti u patsientov s klinicheskim diagnozom semeinoi giperkholesterinemii.

UNLABELLED: Low density lipoprotein receptor (LDLR) gene mutations cause familial hypercholesterolemia which is associated with elevated risk of ischemic heart disease. AIM: To define LDLR gene mutations in unrelated patients with heterozygous familial hypercholesterolemia in Russia. METHODS: PCR- single-strand conformation polymorphism analysis, automated DNA sequencing, and test for the presence of the apolipoprotein (apo) B-3500 mutation known to induce hereditary defect in apo-B-100. RESULTS: We found 6 novel mutations of LDLR gene designated E8X, 230insG, 671_679dupGACAAATCT, W422R, D461Y, and V698L. We also identified three missense mutations - C139G, E207K and R395W, which were previously described in FH patients from western populations. None of the studied persons had apo-B-3500 mutation. CONCLUSION: These findings broaden knowledge on mutations responsible for development of familial hypercholesterolemia and confirm molecular heterogeneity of this disease in Russia.