Principles and Problems of Exosome Isolation from Biological Fluids.

Exosomes, the subclass of small membrane extracellular vesicles, have great diagnostic and therapeutic potential, but the lack of standardized methods for their efficient isolation and analysis limits the introduction of exosomal technologies into clinical practice. This review discusses the problems associated with the isolation of exosomes from biological fluids, as well as the principles of traditional and alternative methods of isolation. The aim of the presented review is to illustrate the variety of approaches based on the physical and biochemical properties of exosomes that can be used for exosome isolation. The advantages and disadvantages of different methods are discussed.

Changes of infection of the prostate tissue by Herpesviridae family viruses and HPV in prostate cancer patients depending on the stage of the pathological process.

The role of infection in the beginning and progression of prostate cancer is not known to the end at the moment. Experimental data, presented in the article, are obtained by hybridization in situ and demonstrate high degree of infection of the prostate tissue by viruses with the oncogenic and oncomodulation properties HCMV, EBV, HHV8 and HPV. The authors revealed that in patients at different substages of T2 stage infection of the prostate tissue increased from T2a to T2c. The importance of these findings is discussed.

[Infection of the prostate by herpesviruses and HPV in patients with prostate cancer].

The exploration of the presence of the HCMV, EBV, HHV8 and HPV-infection in the prostatic tissue of the prostate cancer patients was conducted. The methods of PCR and in situ hybridization (ISH) were used and the results were compared. Authors revealed high degree of virus infection in prostate. Using of PCR and ISH resulted in following frequencies of the viruses in prostate tissue: HCMV-87 and 77%, EBV-53 and 54%, HHV8--0 and 50%, HPV--0 and 23%. The question of the nature of discrepancies is discussed.

[Poorly differentiated prostate cancer may be associated with cytomegalovirus infection in prostatic tissue].

Polymerase chain-reaction procedure was used to test archival samples of prostate tissue from 90 cancer patients for cytomegalovirus infection detected in 46.7%. The median Gleason score and G-index (TNM classification) were significantly higher in CMV-positive patients (8 and 4, respectively) than those in CMV-negative ones (6 and 2, respectively). A relationship between cytomegalovirus infection and poor prognosis of prostate cancer is suggested.

[The experience of diagnosis, treatment and prophylaxis of gastric and duodenal ulcer in outpatient conditions].

Gastric and duodenal ulcer (peptic ulcer) takes the leading place among the pathology of digestive organs. According to the data of specialists from Consulting-and-Diagnostic Center N 52 among the servicemen with chronic diseases the digestive pathology constitutes 32%. Among these diseases the peptic ulcer occurs in 81% cases. Screening esophagogastroduodenoscopy conducted in 1100 servicemen without any complaints aged 22-50 has revealed duodenal ulcer in 5.4%, gastric ulcer--in 0.7% and gastroduodenitis--in 11.4%. With introduction of modern methods of diagnosis, treatment, rehabilitation and prophylaxis of peptic ulcer 98% patients with disease uncomplicated course began and completed treatment in outpatient conditions.

[Gene expression profiles of protein kinases and phosphatases obtained by hybridization with cDNA arrays: molecular portrait of human prostate carcinoma]. / Profili ékspressii proteinkinaz i fosfataz, poluchennye metodom gibridizatsii s uporiadochennymi naborami kDNK (cDNA arrays): molekuliarnyi portret raka predstatel'noi zhelezy cheloveka.

Hybridization with cDNA arrays was used to obtain expression profiles of 263 protein-tyrosine kinase (PTK), protein-tyrosine phosphatase (PTP), dual-specific phosphatase (DuSP), and other genes for the normal prostate tissue, primary prostate carcinomas (PC) of 84 patients, 7 xenografts, and 5 carcinoma cell lines. Analysis of 96 profiles revealed eight clusters of genes coexpressed in PC (coefficient of correlation r > 0.7). According to the known functions of their genes, the clusters were designated as proliferating-cell (CDC42, TOP2A, FGFR3, MYC, etc.), neoangiogenesis and blood-cell (LCK, VAV1, KDR, VEGF, MMP9, SYK, PTPRS, and FLT4), invasion-1 and invasion-2 (ADAM17, TRPM2, DUSP6, VIM, CAV1, CAV2, JAK1, PTPNS1, FYN, and PDGFB), HER2, and PSA/PSM/HER3. Basing on expression profiles of 66 genes, a molecular classification of PC was constructed and allowed discrimination between PC and cell lines or xenografts at 98.9% probability. The results suggested that, along with PSA, PSM (FOLH1), kallikrein-2, and a-2-macroglobulin, cell signaling genes EGFR, HER2, HER3, TOP2, KRT8, KRT18, VEGF, CD44, VIM, CAV1, and CAV2 may serve as diagnostic and prognostic markers in PC. The HER2, VEGF, and CD44 genes and the MMP and ADAM families were assumed to be promising targets for inhibitors of PC cell proliferation and metastasis.

[Molecular portrait of human kidney carcinomas: the gene expression profiling of protein-tyrosine kinases and tyrosine phosphatases which controlled regulatory signals in the cells]. / Molekuliarnyi portret kartsinom pochki cheloveka, poluchennyi na osnove ékspressii protein-tirozin-kinaz i tirozin-fosfataz, kontroliruiushchikh peredachu reguliatornykh signalov v kletkakh.

Hybridization with cDNA arrays was used to obtain expression profiles of 214 protein-tyrosine kinase, protein-tyrosine phosphatase, dual-specific phosphatase, and other genes for kidney carcinomas (KC) and normal kidney tissues of 34 patients and for seven carcinoma cell lines. Computer analysis revealed three clusters of genes coexpressed in KC. A proliferating-cell gene cluster included MET, VIM, MYC, TOP2A, PCNA, etc. A neoangiogenesis and blood-cell gene cluster included LCK, HCK, FGR, MMP9, CSFR1, VEGF, FLT1, and KDR. A cluster corresponding to normal, differentiated kidney cells included ERBB2 (HER2) for receptor protein-tyrosine kinase, several phosphatase genes (PTPRE, PTPRB, DUSP9), and EGF. The results suggested that MET, DUSP9, PCNA, TOP2A, and VIM may serve as diagnostic and prognostic markers in KC. Tubulin and topoisomerase II were assumed to be promising targets for cell proliferation inhibitors in KC.

[Cloning of genes activated in murine embryonal stem cells during differentiation in suspension]. / Klonirovanie geov, aktiviruemykh v émbrional'nykh stvolovykh kletakh myshi v protsesse differentsirovki v suspenzii.

Embryonic stem cells (ESC) are widely employed as an experimental model owing to the similarity of molecular events occurring in their in vitro differentiation and in early embryo development. Subtraction hybridization was used to clone genes that display enhanced transcription in simple embryoid bodies (EB), which are formed on ESC culturing in suspension and are characterized by the presence of endodermal cells. The cloned sequences proved to include three new genes lacking homologs in databases. Northern analysis of the transcript tissue distribution in adult mice confirmed higher expression of these genes in differentiated cells compared with ESC.

Gamma-radiation-induced cell death in the fetal rat brain possesses molecular characteristics of apoptosis and is associated with specific messenger RNA elevations.

Low-dose ionizing irradiation of 16-18-day pregnant rats rapidly kills stem cells in the fetal forebrain. We have examined gamma-irradiated 17-day fetal rat brain tissue for molecular characteristics of apoptosis and changes in levels of mRNAs relevant to apoptosis. In many forebrain cells radiation elicits within 5 h nuclear condensation and fragmentation consistent with apoptosis. An electrophoretic DNA ladder indicative of internucleosomal chromatin cleavage was prominent within 3 h after irradiation. Pretreatment of pregnant rats with cycloheximide, or pretreatment of dissociated fetal brain cells in culture with actinomycin D, abolished the radiation-induced internucleosomal DNA fragmentation, demonstrating requirements for protein and RNA synthesis. Irradiation dramatically increased the level of the p53 transcription factor and the abundances of mRNAs coding for the cell-cycle inhibitor p21/Waf-1/Cip-1 and the AP-1-associated transcription factors Fos and JunB. Irradiation moderately increased the level of mRNA for the positive apoptosis regulator Bax. In contrast, irradiation reduced by 50-70% the abundances of most other mRNAs tested, including those for housekeeping proteins, p53, Jun, Myc, interleukin-1-beta-converting enzyme, and the negative apoptosis regulators Bcl-2 and Bcl-xL. These results indicate that radiation-elicited apoptosis of fetal brain cells is associated with activation of the p53 system, probable increases in AP-1 Fos/JunB heterodimers, and an increased ratio of Bax to Bcl-2 + Bcl-xL.

[Cloning in the lambda ZAPII/pBluescript system. I. Design of common cDNA libraries]. / Klonirovanie v sisteme lambda ZAPPII/pBluescript. I. Konstruirovanie obychnykh konotek kDNK.

Here in I summarize some cloning tips that have been revealed during construction of cDNA libraries from rat tissues and mouse stem cell lines in lambda ZAPII, and that improve the cloning efficiency. These improvements can be applied to many different vectors and cDNA cloning systems.

[Effect of prenatal neutron irradiation on gene expression in the developing rat brain]. / Deistvie prenatal'nogo neitronnogo oblucheniia na ékspressiiu genov v razvivaiushchemsia mozge krysy.

.5 Gy prenatal neutron irradiation at the 17 day of gestation resulted in activation of gene expression in brain. Elevated level of neuronal (GAP-43, NCAM, protein kinase C, calmodulin) and protooncogene (c-foc, c-jun, c-mas) gene transcripts in brain of 2-4 weeks old rats well correlated both with brain maturation in normal animals and development of brain weight deficiency in the irradiated rats. Activation of gene expression appears to be a compensatory response of surviving brain cells to irradiation. Suppression of the gene activity in brain of 18 months old rats correlated with impairment of brain functions at later periods after prenatal irradiation.