In Vivo DYSF Gene Viral Delivery Provides a Histoprotective Effect in Skeletal Muscle Tissue in Dysferlin-Deficient Mice.

We studied the effects of a dual-vector DYSF gene delivery system based on adeno-associated virus serotype 9 capsids on pathological manifestations of dysferlinopathy in skeletal muscles of Bla/J mice lacking DYSF expression. The mice received intravenous injection of 3×1013 genomic copies of the virus containing the dual-vector system. M. gastrocnemius, m. psoas major, m. vastus lateralis, and m. gluteus superficialis were isolated for histological examination in 3, 6, and 12 weeks after treatment. Healthy wild-type (C57BL/6) mice served as positive control and were sacrificed 3 weeks after injection of 150 µl of 0.9% NaCl into the caudal vein. To detect dysferlin in muscle cryosections, immunohistochemical analysis with diagnostic antibodies was performed; paraffin sections were stained with hematoxylin and eosin for morphometric analysis. After administration of gene-therapeutic constructs, muscle fibers with membrane or cytoplasmic dysferlin location were detected in all examined muscles. The proportion of necrotic muscle fibers decreased, the number of muscle fibers with central location of the nucleus increased, and the mean cross-section area of the muscle fibers decreased.

[Association of a polymorphic marker Trp719Arg of KIF6 gene with effects of atorvastatin and simvastatin in patients with early ischemic heart disease].

Action of statins is characterized by pronounced variability what is caused by effects of a multitude of factors. Main of these factors appears to be genetic peculiarity of patients. We studied influence of polymorphic marker Trp719Arg of KIF6 gene on lipid and nonlipid effects of atorvastatin and simvastatin. The studied genetic marker is associated with risk of development of ischemic heart disease and myocardial infarction as well as efficacy of therapy with statins according to data of a number of large multicenter studies. We examined 60 men with ischemic heart disease which had manifested in young age when genetic factors were most expressed and had special significance. Efficacy of 40 mg/day simvastatin did not depend on genotypes of polymorphic marker Trp719Arg of KIF6. Therapy with 10 mg/day atorvastatin was more effective in carriers of polymorphic marker Trp719Arg of KIF6 gene by action on dynamics of changes of high sensitivity C-reactive protein and dispersion of high density lipoprotein response. Increase of atorvastatin dose to 80 mg/day abolished influence of genotypes. Thus for the first time we discovered influence of polymorphic marker Trp719Arg of KIF6 gene on individual response to therapy with 10 mg/day of atorvastatin, while and apoA1, structural protein of high density lipoproteins can be considered as a marker of "fast response".

[Polymorphic markers Ala455Val of the THBD gene and Arg353Gln of the F7 gene and association with unfavorable outcomes of coronary atherosclerosis in patients with a history of acute ischemic heart disease].

The polymorphic markers Ala455Val of the THBD gene and Arg353Gln of the F7 gene were tested for association with the frequency of unfavorable outcomes in patients with a history of acute ischemic heart disease. The study involved 1145 patients hospitalized in cardiology clinics of Moscow, St. Petersburg, Kazan, Chelyabinsk, Perm, Stavropol, and Rostov-on-Don because of acute ischemic heart disease. The patients were followed up for up to 62.5 months. None of the markers displayed a significant association with the time to an endpoint. The patients were then grouped by sex. In females, the frequency of unfavorable outcomes (fatal or nonfatal myocardial infarction and fatal or nonfatal stroke) was higher in carriers of allele Val of the Ala344Val polymorphic marker of the THBD gene and carriers of genotype Arg/Arg of the Arg353Gln polymorphic marker of the F7 gene, but the difference was not statistically significant. Such an increase in frequency was not observed in males. To study the combined effect of the polymorphic markers of the THBD and F7 genes, the course of ischemic heart disease was compared for two female subgroups. One included carriers of allele Val of the Ala344Val polymorphic marker of the THBD gene and genotype Arg/Arg of the Arg353Gln polymorphic marker of the F7 gene; the other subgroup included carriers ofgenotype Ala/Ala of the Ala455Val polymorphic marker of the THBD gene and allele Gln of the Arg353Gln polymorphic marker of the F7 gene. The frequency of unfavorable outcomes in the first subgroup was higher than in the second one. The time to an endpoin was 40.5 months (95% confidence interval (CI) 33.5-47.6) in the first subgroup and 51.6 months (95% CI 45.0-58.1) in the second subgroup (chi2 = 4.15, P = 0.042). The results made it possible to assume that the F7 and THBD genes play an important role in genetic predisposition to unfavorable outcomes in patients with a history of acute ischemic heart disease.

[Gene IL6 G(-174)C and gene IL10 G(-1082)A polymorphisms are associated with unfavourable outcomes in patients with acute coronary syndrome].

We investigated the association of gene IL6 G(-174)C polymorphism and gene IL10 G(-1082)A polymorphism with coronary artery disease (CAD) in the Russian population. A total of 1145 patients with CAD diagnose on the basis of clinical studies in cardiological hospitals of Moscow, St -Petersburg, Kazan, Chelyabinsk, Perm, Stavropol and Rostov-on-Don. Supervision term was 9.10 +/- 5.03 months (the maximum term 18 months). In case of gene IL10 G(-1082)A polymorphism we determined that patients with CAD diagnose and A alleles gene IL10 had unfavorable outcome more often than patients with homozygous G alleles. Survival time from end point from carrier genotype GA and AA is 11.68 +/- 0.67 months against 12.69 +/- 0.65 months from carrier phenotype GG gene IL10 (chi2 = 4.13, p = 0.042). The group studied do not differ significantly with respect to the distributions of gene IL6 G(-174)C alleles and genotypes. However in case combined group studies of gene IL10 G(-1082)A polymorphism and IL6 G(-174)C polymorphism we determined that patients with CAD diagnose and carrier genotype GG gene IL6 and genotype GA and AA gene IL10 had unfavorable outcome more often (survival time 11.01 +/- 1.24 months) than patients with genotype CC and CG gene IL6 and genotype GG gene IL10 (survival time 13.28 +/- 0.83 months) chi2 = 10.23, p = 0.017. The obtained data allows assuming the important role of the IL6 and IL10 genes which are responsible for functioning of inflammation system, in the accelerated formation of failures at the patients who had a coronary syndrome.

[The use of DYS14 marker for sex determination].

The possibilities of real-time PCR amplification of DYS14 marker located on Y chromosome for sex determination were studied. Samples of plasma of 30 men and 30 women were investigated for this aim. Real-time PCR amplification of DYS14 marker (located inside gene coding TSPY1 protein) was used for sex determination. According to the obtained results, 30 samples belonged to men and 30--to women. In all our experiments the results were confirmed by use of marker SRY, widely used in forensic examination. Detection limit of DNA region containing DYS14 in reaction mixture was established after experiment with dilution of male DNA and is equal to 6.7 pg of DNA (two copies of genome), which corresponds to 6.7 ng of DNA (2000 copies of genome) in 1 ml of blood. Sex determination with small amounts of genetic material in investigated sample becomes possible with such characteristics. Method can be used for noninvasive prenatal diagnostics for the timely detection of congenital diseases associated with sex and in forensic medical examination.