[Comparative transcriptome analysis of human aorta atherosclerotic lesions and peripheral blood leukocytes from essential hypertension patients].

One of the major cardiovascular risk factor which predisposes to and accelerates atherosclerosis is arterial hypertension (AH). To determine the molecular basis of the crosslink between AH and atherosclerosis for the development of new treatment strategies large-scale transcriptome analysis of the cells implicated in atherogenesis is needed. We used cDNA microarray technique for simultaneous analysis of gene expression in human abdominal aorta normal sites and atherosclerotic lesions of different histological types, as well as in peripheral blood leukocytes from patients with essential hypertension (EH) and donors. The microarray data were verified by quantitative RT-PCR (reverse transcription coupled with polymerase chain reaction) and immunohistochemical analysis. Differential expression of 40 genes has been found, among which twenty two genes demonstrated up-regulation and 18 genes demonstrated down-regulation in atherosclerotic aorta compared with normal vessel. New gene-candidates, implicated in atherogenesis, have been identified - FPRL2, CD37, CD53, RGS1, LCP1, SPI1, CTSA, EPAS1, FHL1, GEM, RHOB, SPARCL1, ITGA8, PLN, and COL14A1. These genes participate in cell migration and adhesion, phenotypic changes of smooth muscle cells, immune and inflammatory reactions, oxidative processes and extracellular matrix remodeling. We have found increased expression levels of CD53, SPI1, FPRL2, SPP1, CTSD, ACP5, LCP1, CTSA and LIPA genes in peripheral blood leukocytes from EH patients and in atherosclerotic lesions of human aorta. The majority of these genes significantly (p<0.005) positively (r>0.5) correlated with AH stage as well as with histological grading of atherosclerotic lesions.

[Gene expression analysis in myocytes of right atrial appendages in patients with atrial fibrillation using cDNA microarray technique].

Gene expression level of 2900 genes was studied by cDNA microarray in patients with atrial fibril-lation (AF) or sinus rhythm. Gene transcripts were analysed in samples of right atrial appendages from 47 patients undergoing surgery for valve repair or coronary artery bypass. Standard correlation analysis and two dimensional hierarchical clustering were used for study of differentially expressed genes in patient groups. A highly positive correlation of gene expression with AF was shown for cardiac muscle LIM domain protein (CSRP3), cardiac muscle myosin heavy chain beta isoform (MYH7), calmodulin (CALMS) and homeobox protein (PKNOXl) genes (r > 0.77, p < 0.007). In contrast, metallothionein (MT1/2), mitochondrial aldehyde dehydrogenase 2 (ALDH2), ras-related protein (RaplA) and guanine nucleotide binding protein G (GNAL) genes revealed highly negative correlation with AF (r < -0.75, p < 0.002). Alterations of gene activity were more evident at permanent as compared with paroxysmal AF. In addition, genes overexpressed in AF patients demonstrated underexpression in coronary artery disease patients (r=-0.8, p=0.0002) and conversely. Genes correlating with AF belong to different functional categories, including sarcomere organization, contraction, Ca2+ homeostasis, signaling and transcription regulation, extracellular matrix interactions and oxidative stress. Downregulation of MT1/2 and ALDH2 genes, known protectors against oxidative stress, may contribute to maintenance of oxidative stress in myocardial tissues of AF patients. The identification of novel genes - participants of pathological process in AF may open new perspective for search of therapeutic agents.

[Phospholipase A2, group IIa, as an earlier unknown immunohistological marker of cardiac myxoma].

Phospholipase A2, group IIA, gene expression has been analyzed in primary heart tumors. High expression has been demonstrated through several ways: reverse-transcriptase chain polymerase chain, Northern blotting hybridization at the RNA level and immunoblotting, immunohistochemical assay at the protein level. Human cardiac myxoma exhibits highly positive phospholipase A2, group IIA, immunophenotype (100% positive cases). The immunophenotype is unique among human primary cardiac tumors. Phospholipase A2, group IIA, can be proposed as a tissue marker for pathological examination after heart tumor resection.

[PRKAR1A gene mutations in two patients with myxoma syndrome (Carney complex)]. / Mutatsii v gene reguliatornoi sub"edinitsy 1-alpha tsiklo-AMF-zavisimoi proteinkinazy A (PRKAR1A) u dvukh bol'nykh s miksomnym sindromom (kompleksom Carney).

Carney complex is an autosomic dominant disorder initially described as the association of cardiac myxomas, spotty skin pigmentation and endocrine overactivity and considered as a multiple neoplasia and lentiginosis syndrome. Mutations in the tumor suppressor gene PRKAR1A, coding for the type 1-alpha regulatory subunit of cAMP-depended protein kinase A have been previously identified in about half of the Carney complex kindreds. In this paper we report identification of the molecular defect in PRKARIA gene in two Carney complex patients. A new mutation (403delAC) located in a 3rd exon of PRKARIA gene has been observed in one case, and a previously described mutation in exon 7 (847delTC) in the second case.

[mRNA for the plasminogen activator of the urokinase type: translation in oocytes of Xenopus laevis]. / mRNK aktivatora plazminogena urokinaznogo tipa: transliatsiia v ootsitakh Xenopus laevis.

Poly(A)+-RNA from human kidney and human embryonal lung fibroblasts fractionated by sucrose gradient centrifugation was translated in Xenopus oocytes. Assay for plasminogen-dependent fibrinolytic activity detected synthesis of secreted plasminogen activator and revealed the active fraction of poly(A)+-RNA with a sedimentation coefficient of approximately 23S. Translation products of the active fraction were immunoadsorbed by antiurokinase monoclonal antibodies immobilized on sepharose. Gel electrophoretic analysis of the protein products showed that the 23S fraction of poly(A)+-RNA from human kidney contains mRNA for single-chain urokinase-type plasminogen activator with apparent molecular weight of approximately 50 kDa.

[Non-specific RNA degradation in the presence of magnesium ions]. / Nespetsificheskaia degradatsiia RNK v prisutstvii ionov magniia.

RNA degradation catalyzed by Mg2+ was studied under the conditions of reverse transcriptase reaction. Agarose gel electrophoresis in 6 M urea was employed to follow the reaction. Natural ribosomal of poly(A)+ RNA as well as synthetic poly(rA) are equally accessible for degradation. Neither an RNAase nor alkali alone is responsible for the degradation. The reaction rate is directly proportional to Mg2+ concentration in the range of 1 to 10 mM, doesn't depend upon RNA concentration and enhances approximately 50 fold upon increasing of pH value from 7.5 to 9.0. It was concluded that the Mg2+ catalyzed degradation reaction is an unspecific one in respect to the primary structure of RNA. The results obtained are useful to optimize the conditions for the reverse transcriptase reaction.

[Isolation and characteristics of urokinase-type plasminogen activator from a culture of human embryo lung fibroblasts]. / Vydelenie i kharakteristika aktivatora plazminogena urokinaznogo tipa iz kul'tury legochnykh fibroblastov émbriona cheloveka.

Plasminogen activator from conditioned medium of human embryonal lung fibroblasts was purified by phosphocellulose P11 chromatography, followed by p-aminobenzamidine-agarose chromatography. Two forms of plasminogen activators were separated by chromatography on the heparin-sepharose. The high molecular weight form (53 kDa) with specific activity 130 000 IU/mg consists of two polypeptide chains (31 kDa and 20 kDa) and exhibits strong affinity for fibrin-celite, lysine-sepharose and heparin-sepharose. The low molecular weight form (32 kDa, 190 000 IU/mg) also binds to these sorbents, but more weakly, and its properties are very similar to those of low molecular weight urokinase. Activity of both forms of plasminogen activators are inhibited by monoclonal antibodies against urokinase. A number of enzymological chromatographic and immunological properties indicates, that the plasminogen activator from lung fibroblasts is of urokinase type.

[Isolation and characteristics of the 8--10S poly(A)-containing mRNA fraction from mid-blastula stage Stronglyocentrotus intermedius sea urchin embryos]. / Vydelenie i kharakteristika fraktsii 8--10S poli(A)-soderzhashchikh mRNK iz émbrionov morskogo ezha Strongylocentrotus intermedius na stadii srednei blastuly.

Centrifugation in sucrose linear gradient and affinity chromatography on oligo (dT)-cellulose were used to isolate the poly(A)-containing fraction of 8--10S RNA from St. intermedius sea urchin embryos at the middle blastula stage. This RNA has been shown to possess template activity in the cell-free protein-synthesizing system from wheat germs. The studied mRNA fraction induces synthesis of two polypeptides with the molecular weights of 10 and 15 thousand daltons according to the data of electrophoresis in polyacrylamide gel in the presence of sodium dodecyl sulphate. The products of in vitro translation were immunochemically identified with embryo-specific glycoproteins of the plasma membrane of embryo cells.

[Translation of MOPC 21 plasmacytoma cell mRNA in a cell-free wheat germ system]. / Transliatsiia v beskletochnoi sisteme iz zarodyshei pshenitsy mRNK kletok plazmotsitomy MOPC 21.

A wheat germ cell-free system was used for translation of kappa-chain IgG1 messenger RNA isolated from mouse plasmacytoma MOPC 21 cells polysomes. The system was optimised for a number of ingredients of the incubation mixture. Incorporation of labelled amino acids in polypeptides was shown as a function of K+, Mg2+, exogenous mRNA concentration and time, mRNA was purificated by two successive oligo (dT)-cellulose chromatographies and two successive sucrose gradient centrifugations or polyacrylamide gel electrophoresis. Different fractions of poly(A)-RNA stimulated protein synthesis 20--30-fold in a wheat germ cell-free system. Analysis of the translation product of 14 and 16S mRNA by SDS-polyacrylamide gel electrophoresis revealed polypeptide comigrating with the authentic L-chain IgG1 and specifically precipitating by an antiserum to mouse IgG and L-chain IgGl (MOPC 21). The 18 and 28S mRNA fractions directed the synthesis of polypeptides with molecular weight up to 50 000 dalton. Immunospecific translation product of the 18 and 28S mRNA contain both L- and H-chains.

[Isolation and characterization of the matrix properties of electrophoretically homogeneous immunoglobulin L-chain mRNA from mouse myeloma MORC 21]. / Vydelenie i kharakteristika matrichnykh svoistv élektroforeticheski gomogennoi mRNK L-tsepei immunoglobulina mielomy myshei MORC 21.

mRNA with m. w. 4.10(5) dalton was isolated by elution of one of the fractions of poly(A)-RNA from polyacrylamide gel. During reelectrophoresis this mRNA was electrophoretically homogeneous and had the same size. The isolated fraction represented the template activity in a wheat germ cell-free system of protein synthesis and stimulated the synthesis of polypeptides, electrophoretically correspondent to L-chain of authentic immunoglobulin. Comparative data of properties of electrophoretically homogenous mRNA and analogous fraction of mRNA, isolated from sucrose concentration gradient after fractionation of total poly(A)-containing RNA are given.

[mRNA of mouse plasmacytoma. Reverse transcription and translation in cell-free systems]. / Matrichnye RNK plazmotsitony myshei. Obratnaia transkriptsiia i transliatsiia v beskletochnoi sisteme sinteza.

Total poly(A)-mRNA from polyribosomes of MOPC 21 mouse myeloma were investigated. Poly(A)-mRNA was released by two successive chromatography on oligo (dT)-cellulose. A 14S fraction of total poly(A)-mRNA was obtained and partially purified by sucrose gradient centrifigation followed by acrylamide gel electrophoresis. As estimated from the electrophoretic analysis, the 14S mRNA has three components, one of which appears to be 18S rRNA and two others--mRNAs with molecular weight of 5.2.10(5) and 3.8.10(5), respectively. Total poly(A)-mRNA and partially purified 14S mRNA were active when employed as a template in a reverse transcription and cell-free system from wheat germ. DNA complementary to the 14S mRNA was prepared with avian myeloblastosis virus RNA-dependent DNA polymerase. This cDNA was heterogeneous in size with the average size of about 800 nucleotides when analyzed by gel electrophoresis in 98% formamide. The maximal length was about 1100 nucleotides that consistent with full template length. About half of the translation product directed by the 14S mRNA migrated as mature L-chain Ig (upon polyacrylamide gel electrophoresis in sodium dodecylsulfate). The presented data suggested that 14S mRNA species contain mRNA L-chain Ig.

[mRNA fractions of mouse plasmacytoma cells]. / Fraktsii mRNK kletok plazmatsitomy myshei

The sedimentation characteristics of polysomal mRNA labelled in vitro by [5-3H]uridine and electrophoretic mobility of similar non-labelled mRNA of mouse plasmacytoma were studied. Rapidly labelled polysomal mRNA may be considered as mRNA on the basis of several independent but indirect tests. These mRNA's are localized in 18-6S region of sucrose gradient. Some part of radioactivity have been found in the ribosomal RNA. It was shown that there is 8-10 RNA fractions in sucrose gradient. The 16S and 12-14S fractions are isolated and partially purified by two- and three-fold centrifugation. Fractions homogenous in sucrose gradient were electrophoresed in PAAG-SDS and divided into several subfractions some of them being common for 16S and 12-14S. The number of non-crossover subfractions was about 2-3. Not less than 20 different main fractions of polysomal mRNA were determined in plasmacytoma cells on the basis of electrophoretic data.