[Autoimmune hemolytic anemia associated with mesenteric teratoma]. / Autoimmunnaia gemoliticheskaia anemiia, assotsiirovannaia s teratomoi bryzheiki tonkoi kishki.

The paper describes a case of autoimmune hemolytic anemia (AIHA) in a 27-year-old woman whose examination revealed mesenteric teratoma. AIHA was characterized by a hypertensive crisis and a temporary response to corticosteroid therapy that was complicated by the development of somatogenic psychosis and discontinued. A relapse of hemolysis developed 6 months later. The patient underwent laparoscopic splenectomy and removal of mesenteric root teratoma. Immediately after surgery, a hematological response was obtained as relief of hemolysis and restoration of a normal hemoglobin level. There is a sustained remission of AIHA for the next 16 months.

[Comparative Analysis of Polymorphisms of the Serotonin Receptor Genes HTR1A, HTR2A, and HTR1B in Hadza and Datoga Males].

The distribution of the allele and genotype frequencies of polymorphic loci of serotonin receptor genes (HTR1A, rs6295; HTR2A, rs6311; HTR1B, rs6296) in Hadza (n = 197) and Datoga males (n = 230) living in Tanzania was determined. It was shown that the populations significantly differ by the frequencies of alleles and genotypes of the rs6295 locus of the HTR1A gene. The G-allele (0.779) and the genotype G/G (0.590), which are markers of increased risk of suicidal and impulsive behavior, respectively, are revealed in Hadza with high frequency. It was found that the frequency of homozygous G/G of the rs6296 locus of the HTR1B gene, which is a marker of increased risk of outward directed aggression, is higher in Datoga (0.563) than in Hadza (0.457). The allele and genotype frequencies of the rs6311 locus of the HTR2A gene do not differ among the Hadza and Datoga males. The data on the distribution of allele and genotype frequencies of the HTR1A, HTR2A, and HTR1B genes can be used to determine the associations of the identified markers with various forms of human aggressive behavior.

[The 3'-UTR polymorphism of dopamine transporter gene in hadza and datoga males].

The study of VNTR-polymorphism and the molecular struc ture of 3'-UTR of the dopamine transporter gene (DAT1/SLC6A3) was performed in hadza and datoga males. It was shown that hadza and datoga differed in allele and genotype frequencies. Allele with 9 repeats in 3'-UTR is more common in hadza as well as homozygous genotype DAT19/9. Allele with 10 repeats is more common in datoga as well as homozygous genotype DAT1 10/10. The molecular structure of the DAT alleles with 3, 8 and 12 repeats was determined for the first time. In addition it was found that DAT1 allele with 11 repeats in datoga significantly differed from previously described ones in other populations in repeats type and arrangement. We suggest that variations of the repeats num ber and type in the 3'-UTR of allelic variants may affect the dopamine transporter gene function.

[Clinical significance of insulin resistance syndrome in development of endothelial dysfunction in patients with primary gout].

Endothelial function has been studied in 175 males with primary gout. It has been established that incidence of endothelial dysfunction in patients with primary gout was associated with the clinical course and was marked most of all in patients who had a chronic gout. The correlations found between the parameters of endothelial dysfunction and impaired carbohydrate metabolism suggest close link and coordination between the above processes which contribute greatly to development and progress of atherosclerosis in such patients.

[Clinical significance of abnormal fatty acid composition in red cell membranes and carbohydrate metabolism in patients with primary gout and arterial hypertension].

We studied changes in fatty acid composition in the lipid fraction of red cell membranes and carbohydrate metabolism in patients presenting with primary gout with and without arterial hypertension. They were characterized by an increased content of saturated and decreased level of unsaturated fatty acids. The latter contained more monoenoic and significantly less polyenoic acids compared with the normal values. The pool of polyunsaturated fatty acids (PUFA) contained a small amount of w-3 PUFA whereas that of w-6 PUFA varied oppositely, i.e. the relative amount of gamma-linolenic acid was elevated and that of dihomo-gamma-linolenic acid significantly decreased compared with healthy men. The fractional composition of higher fatty acids in patients with gout depended on AH and peculiarities of carbohydrate metabolism. The data obtained suggest the contribution of disturbances in fatty acid metabolism and insulin resistance to the development of AH syndrome in patients with primary gout.

[Peptide inhibitors of myosin light chain kinase. Development of peptidase resistant analogues].

Myosin light chain kinase (MLCK) is the key regulator of various forms of cell motility including endothelial and epithelial permeability in particular. One of the potential MLCK inhibitors to be used in humans is a membrane permeable peptide H-RKKYKYRRK-NH2 (L-PIK). In present work we used solid phase peptide synthesis and Fmoc-technology to produce five modifications of L-PIK. Based on (1)H NMR analysis revealed that these peptides demonstrated improved resistance to degradation in blood plasma. One of de novo synthesized peptides, L-[MeArg(1)]PIK inhibited MLCK activity in vitro with the same efficiency as L-PIK whereas other modified peptides showed reduced inhibitory activity. D-amino acid analog of PIK was the least active inhibitor. Thus, we have demonstrated the possibility to produce an effective MLCK peptide inhibitor with increased resistance to biodegradation that is suitable for further pharmacological development.

[Immunological similarity of diphtheria toxin and EGF receptor].

Cardiomyopathy and neuropathy are the two commonly observed complications in diphtheria patients and in, some instances, individuals vaccinated against diphtheria. The nature of these complications remains not well understood. It was suggested that autoimmunity may play a role in the development of these afflictions. Based on functional similarities between diphtheria toxin (DT) and epidermal growth factor receptor (EGFR), which both can bind to the heparin-binding EGF-like growth factor (HB-EGF) precursors, we suggested that antibodies developed against DT can cross react with EGFR. Here, using serum from healthy donors (n = 10) and diphtheria patients (n = 15), we demonstrated that B-subunit of DT has the antigenic epitopes similar to those of EGFR. Diphtheria toxin as well as EGFR could be recognized by antibodies raised against EGFR and by serum antibodies from diphtheria patients. Moreover serum of diphtheria patients competitively inhibits binding of anti-EGFR antibodies to the receptor. The truncated diphtheria toxin without B-subunit could be detected by serum antibodies of diphtheria patients, but not by anti-EGFR antibodies. Collectively, these studies demonstrate cross-reactivity of antibodies raised against B-subunit of DT and extracellular domain of EGFR and suggest that clinically observed post-diphtheria complications may result from autoimmune inhibition of EGFR function and possible destruction of receptor-positive tissues.

[Thromboelastographic characteristics of different infusion therapy regimens in healthy bone marrow donors].

Infusion solutions are able to change the hemostatic system. Thromboelastography (TEG) is an integral technique to evaluate the hemostatic system. TEG was used to evaluate the effect of three infusion solutions (6% hydroxyexyethyl starch (HES) 200/0.5 - Hemohes; HES 130/0.4 - Voluven; modified gelatin solution - Gelofusin) on the hemostatic system in 36 bone marrow donors (healthy individuals). The solutions were used in combination with crystalloid solutions during a procedure to compensate for intraoperative blood loss. Hemostatic changes were noted by the end of an operation in all groups; however, these were less pronounced when Voluven was administered. Thus, all colloid infusion solutions have varying effects on the hemostatic system, with a tendency toward both hypo- and hypercoagulation. According to TEG, HES 130/0.4 (Voluven) has a minimal effect on the hemostatic system.

[Genetic analysis of Drosophila melanogaster chromosome 3 neurodegenerative mutants induced with ethyl methanesulfonate].

Neurodegeneration, a pathological state accompanied by brain neuronal necrosis and changes in behavior, has been described for many animal species. However, the genetic control and molecular mechanisms of this process are yet vague. A large collection of neurodegenerative mutants of a model object, Drosophila melanogaster, can enhance understanding of these mechanisms. In this work, we have demonstrated that genetically determined anatomical changes in Drosophila brain are accompanied by a decreased lifespan and deviations from the wild-type sexual behavior and locomotor activity. It has been found that the genes vacuous and loechrig are candidates for molecular genetic analysis in eight mutants from the collection.

[Enhancement of coagulation caused by transfusion of artificial plasma-replacement solutions].

AIM: To investigate hemostasis disorders caused by massive blood transfusions of artificial plasma replacing solutions (PRS). MATERIAL AND METHODS: Two groups of patients were examined: 7 healthy volunteers without blood loss (group 1) and 11 healthy donors of bone marrow with intraoperative blood loss 1-2 l (group 2). Five patients of group 1 received transfusion of 12 ml/kg hydroxyethyl starch (HES) 130/0.4, two patients of group 1 received transfusion of modified gelatine solution (gelofusin). All of them received infusions (1-1.5 l) of crystalloid PRS (1-2 l) and infusion of one of colloid PRS (6-HES, 5--gelofusin). Estimated hemodilution in group 1 was 1.17 +/- 0.01 times, in group 2 it varied from 1.3 to 2.7 times (mean 1.78 +/- 0.4 times). Hemostasis was studied by clot growth rate (for groups 1 and 2), endogenic thrombin potential and parameters of thromboelastography (for group 2) in plasm samples obtained before, 2.24 and 48 hours after infusion of colloid PRS. RESULTS: For both groups spatial clot growth rate 2 hours after hemodilution was high. Then it fell and reached baseline level 48 hours after PRS infusion. Endogenic thrombin potential and thromboelastography data (for group 2) changed by the same pattern. A hypercoagulation effect of gelofusin on parameters of thromboelastography and clot growth rate was higher than of HES 130/04. CONCLUSION: Moderate hemodilution with PRS in vivo causes hypercoagulation which persisted longer than volemic effect of PRS.

Mechanism of enhancement in electromagnetic properties of MgB2 by Nano SiC doping.

A comparative study of pure, SiC, and C doped MgB2 wires has revealed that the SiC doping allowed C substitution and MgB2 formation to take place simultaneously at low temperatures. C substitution enhances H_{c2}, while the defects, small grain size, and nanoinclusions induced by C incorporation and low-temperature processing are responsible for the improvement in J_{c}. The irreversibility field (H_{irr}) for the SiC doped sample reached the benchmarking value of 10 T at 20 K, exceeding that of NbTi at 4.2 K. This dual reaction model also enables us to predict desirable dopants for enhancing the performance properties of MgB2.

[KRP/telokin differentially regulates filament assembly and phosphorylation of light chains of non-muscle myosin II in fibroblasts].

Transgenic 3T3 fibroblasts have been generated that express either the wild-type KRP or its truncated mutant lacking the C-terminal domain, which primarily contributes to myosin binding of KRP. It was found that KRP-expressing cells display a significantly increased content of myosin filaments and a reduced level of rMLC phosphorylation, whereas the mock transfected cells or cells expressing the C-terminally truncated KRP do not. Our results suggest that (1) KRP promotes the polymerization of myosin II and reduces the rMLC phosphorylation level in cells, (2) KRP acts through direct binding to myosin II, and (3) transgenic 3T3 fibroblasts stably expressing KRP represent a useful and versatile model to study the role of myosin II filament dynamics in cell motility.

[Modified nested polymerase chain reaction in one tube for detection of Enterovirus]. / Modifitsirovannyi metod gnezdovoi polimeraznoi tsepnoi reaktsii v odnoi probirke dlia detektsii énterovirusov.

A method of a modified nest-type polymerase chain reaction (MN-PCR), made in one tube, was elaborated that enhances the sensitivity and cuts the risk of cross-contamination in enteroviruses (EV) detection. The method, as described in detail above, was used to detect EV RNA in 76.9% and 31.25% of examined autopsy samples (13 liquor and 16 cardiac-tissue samples, respectively). It enabled the detection of EV RNA in 6.25% of samples that used to be negative, when tested by MN-PCR in 2 tubes. MN-PCR with one tube is a reliable, sensitive and specific diagnostic tool; it can be recommended for the routine diagnostics of enterovirus infection.

[Incidence of urination disorders and erectile dysfunction after surgical treatment of congenital anorectal defects in children]. / Chastota rasstroistv mocheispuskaniia i érektil'nykh disfunktsii posle khirurgicheskoi korrektsii anorektal'nykh porokov razvitiia u detei.

For assessment of the incidence and characteristics of urination disorders and erectile dysfunction after radical surgical treatment of anorectal congenital defects in children, 60 patients (12 girls, 48 boys) without urination disorders before operation, who have undergone 1-4 proctoplasties were examined. Their was Hirschsprung's disease in 20 patients, anorectal atresia in 40 patients. Iatrogenic urination disorders and erectile dysfunction were revealed in 38 (63.3%) patients, majority of them (24) underwent repeated proctoplasty. The main urination disorders were urinary incontinence (30 patients), infravesical obstruction (10), hyporeflexion of urinary bladder. Erectile dysfunction after operation was in 11 (34.4%) patients from 32. The combination of disorders was revealed in 17 (44.7%) patients. Primary radical operations in children with anorectal defects often cause iatrogenic urination disorders and erectile dysfunction. The number of complications increases progressively after repeated proctoplasty. It is necessary to improve the treatment and the methods of surgical correction bearing in mind the risk of iatrogenic disorders of genitourinary apparatus.

Overexpression of bacterial RecA protein stimulates homologous recombination in somatic mammalian cells.

The pairing of homologous molecules and strand exchange is a key event in homologous recombination promoted by RecA protein in Escherichia coli. Structural homologs of RecA are widely distributed in eukaryotes including mouse and man. As has been shown, human HsRad51 protein is not only structural but also functional homolog of RecA. The question arises whether the bacterial functional homolog of Rad51 can function in mammalian cells and increase the frequency of the homologous recombination. To investigate possible effects of bacterial RecA protein on the frequency of homologous recombination in mammalian cells, the E. coli RecA protein fused with a nuclear location signal from the large T antigen of simian virus 40 was overexpressed in the mouse F9 teratocarcinoma cells. We found that the frequency of gene targeting at the hprt locus was 10-fold increased in the mouse cells expressing the nucleus-targeted RecA protein. Southern blot analysis of individual clones that were generated by targeting recombination revealed predicted type of alterations in hprt gene. The data indicate that the bacterial nucleus-targeted RecA protein can stimulate homologous recombination in mammalian cells.

Camptothecin enhances random integration of transfected DNA into the genome of mammalian cells.

In order to study the involvement of DNA topoisomerase I (top1) in recombination, we examined the effect of the anti-neoplastic drug camptothecin, which selectively poisons top1 by trapping top1-cleavable complexes on integration of exogenic vector into the genome of mammalian cells. We transfected mouse F9 teratocarcinoma cells as well as Chinese hamster V79 cells with a plasmid carrying a selectable neo gene treated with camptothecin, and determined the frequency of neo+ (G418(R)) colonies. We found that treatment with camptothecin for as short a time as 4 h after electroporation resulted in a 4- to 33-fold stimulation of plasmid integration into the recipient genome via non-homologous recombination. These results imply that top1-cleavable complexes trapped by camptothecin could be potentially recombinogenic structures and could stimulate non-homologous recombination in vivo, promoting the integration of transfected plasmids into mammalian genome.

[Hot start of the polymerase chain reaction using DNA helicase]. / Goriachii start polimeraznoi tsepnoi reaktsii pri pomoshchi DNK-helikaz.

A novel method for the hot start of PCR using DNA helicases is developed. The addition of a DNA helicase prevents the random annealing of primers and synthesis of nonspecific products during the preparation of the reaction mixture and initial heating. The hot start of PCR occurs automatically after inactivation of the DNA helicase upon heating of the reaction mixture.

[Effect of inhibitors of topoisomerases and poly(ADP-ribosylation) on homologous and non-homologous integration of exogenous DNA in genes of mammalian somatic cells]. / Vliianie ingibitorov topoizomeraz i pol(ADF-ribozilirovaniia) na na gomologichnuiu i negomologichnuiu integratsiiu ékzogennoi DNK v genom somaticheskikh kletok mlekopitaiushchikh.

A study was made of the influence of inhibitors of poly(ADP-ribose)polymerase, topoisomerase I and topoisomerase II on the frequency of gene targeting of hprt gene as well as on the frequency of random integration of targeting vector pRV9.1 into genome of mouse F9 teratocarcinoma cells. We found that the treatment of cells with the inhibitor of poly(ADP-ribose)polymerase 3-aminobenzamide after electroporation resulted in 3-4-times increase of homologous integration of exogenic vector into chromosomal DNA, and did not affect the frequency of random insertion of transfected DNA. The treatment of cells after electroporation with inhibitors of topoisomerases VP-16, ICRF-193 enhanced random integration of transfected DNA but exerted no effect on the frequency of gene targeting in this experimental system.