CELL CULTURES Information Bulletin, Issue 27, St. Petersburg, 2011. ISSN 2077-6055

CONTENTS T.D. Smirnova, D.M. Danilenko, T.M. Gudkova, M.M. Pisareva, R.A. Kadyirova, A.V. Slita DIFFERENTIAL INFLUENCE OF INFECTIOUS DOSES OF INFLUENZA A VIRUSES ON PROLIFERATION OF HUMAN MONOLAYER CELL CULTURES. pp. 3–12. T.I. Shchelkunova and I.S. Shchelkunov PROPERTIES OF TWO CONTINUOUS CELL LINES ESTABLISHED FROM FIN TISSUE OF SIBERIAN STURGEON, ACIPENSER BAERII. pp. 13–24. E.Y. Adoeva ORGANOTYPIC CULTURING OF EMBRYONIC RAT PANCREAS. pp. 25–33 T.K.Yakovleva, N.M.Yartseva, V.I.Turilova KARYOTYPIC PROGRESSION OF THE HUMAN ACUTE MYELOID LEUKEMIA CELL LINES. pp. 34–45 L.F. Litvinchuk, N.V. Razdolskaya, M.V. Potapchuk, O.V. Gavrilova, A.M. Ivanov BIOLOGICAL PROPERTIES AND CRYOPRESERVATION OF PROTOZOAN PARASITE TRICHOMONAS VAGINALIS. pp. 46–58 V.M. Semenova, L.D. Lyubych, N.I. Lisyany, A.Y. Glavatskiy, L.P. Stayno STUDY OF THE BIOLOGICAL PROPERTIES OF RAT PROGENITOR NEURAL CELLS SUPERNATANT ON BRAIN GLIOMA CULTURES. pp. 58–68 A.V. Lavrov, Y.I. Voldgorn SPECIFICS OF CHROMOSOMES' POSITION IN THE NUCLEUS OF HUMAN MESENCHIMAL STEM CELLS CULTIVATED IN VITRO. pp. 68–73 A.Yu. Markaryan, A.A. Bakharev, P.V. Ustjanzev CELL LINE STANDARDIZATION OPPORTUNITIES BY GENOTYPING WITH SHORT TANDEMIC REPEATS. pp. 74–80 R.Ya. Podchernyaeva, L.V. Uryvaev, A.V. Dedova, L.V. Dedova, K.S. Ionova, G.R. Michailova, N.A. Parasyuk, O.M. Grinkevich, T.K. Selivanova, T.V. Grebenikova, A.D. Petrachev, M.N. Shetvin DETECTION OF MYCOPLASMA AND BOVINE VIRAL DIARRHEA VIRUS CONTAMINATION IN THE COLLECTION OF CELL LINES. pp. 80–88 Instructions to Authors DIFFERENTIAL INFLUENCE OF INFECTIOUS DOSES OF INFLUENZA A VIRUSES ON PROLIFERATION OF HUMAN MONOLAYER CELL CULTURES T.D. Smirnova, D.M. Danilenko, T.M. Gudkova, M.M. Pisareva, R.A. Kadyirova, A.V. Slita Research Institute of Influenza of Ministry of Health and Social Development of Russia, Saint-Petersburg; cellcultures@influenza.spb.ru      Various doses of infection of influenza A viruses A/Brisbane/10/07 and A/S.-Petresburg/56/09 were tested in 8 human monolayer cell lines: ECV-304, T-98G, A-172, RD, CaCo-2, HeLa, L-41, FLECH. The virus-dependent decrease of number of cells grown was registered 72 post-infection and cytopathic changes were observed. When high doses of infection were used the cells responded by induction of apoptosis which was seen earlier than the first signs of cytopathic effect. Only two cell lines - ECV-304 and T-98G - demonstrated enhanced cell proliferation in response to low doses of viral infection and no apoptosis was observed. Detection of viral NP and viral RNA in ECV-304 cells by polymerase chain reaction and fluorescent monoclonal antibodies confirmed that these cells maintained latent influenza A infection for at least 3 serial passages when infected with low doses of A/Brisbane/10/07. Antiviral drugs ribavirine, rimantadine and triazavirine decreased virus-induced proliferation of endothelial cells stimulated by low doses of virus pointing out the specificity of viral influence on the cells. The results of the experiments indicate that the response of a given cell culture is differential and depends on the infection dose of the virus. A possible role between infection of endothelial and neuroglia cells by influenza virus in patients and the subsequent occurrence of cardiovascular pathologies is discussed. Key words:  influenza A viruses, human monolayer cell cultures, proliferation, apoptosis PROPERTIES OF TWO CONTINUOUS CELL LINES ESTABLISHED FROM FIN TISSUE OF SIBERIAN STURGEON, ACIPENSER BAERII T.I. Shchelkunova and I.S. Shchelkunov All Russia Research Institute for Veterinary Virology and Microbiology, Pokrov, Vladimir Province, Russia; shchelkun@yahoo.com      Data on establishment of two cell lines (SSF-1 and SSF-2) from Siberian sturgeon caudal fin tissue, their morphological peculiarities and cultivation parameters, verification of origin and fish virus susceptibility are presented. Key words:  Siberian sturgeon, fin, continuous cell line, characterization ORGANOTYPIC CULTURING OF EMBRYONIC RAT PANCREAS E.Y. Adoeva Military Medical Academy, St. Petersburg, Russia; adoeva@mail.ru      A long-term organotypic culturing of embryonic rat pancreas is carried out. In organ cultures of fetal rat pancreas there is a transformation of acinuses into epithelial tubes. Epitheliocytes of excretory ducts and epithelial tubes are actively dividing and undergo a process of secondary secretory differentiation. Sources of insulocytes in organ cultures of embryonic pancreas: 1) the insulocytes appearance as a part of epithelial tubes, 2) endocrine cells proliferation 3) acino-insular epithelium transformation. Key words:  organ cultures, acinus, insulocyte, secondary differentiation, excretory ducts, the epithelial tubes, acino-insular transformation KARYOTYPIC PROGRESSION OF THE HUMAN ACUTE MYELOID LEUKEMIA CELL LINES T.K.Yakovleva, N.M.Yartseva, V.I.Turilova Institute of Cytology RAS, St.Petersburg, Russia; tyak@mail.cytspb.rssi.ru      Leukemia cell lines are robust and powerful model systems for investigating basic and applied aspects of leukemia cell biology. However, long-term cell cultivation led to further chromosomal rearrangements and karyotypic progression of cell lines. Primary chromosomal abnormalities underlie acute myeloid leukemia (AML) pathogenesis in vivo, but their significance for karyotypic progression of leukemia cell lines during long-term cultivation should be elucidated.      To study the karyotypic variability, the analysis of the published molecular-cytogenetic data of the GF-D8, UoC-M1, HL-60, KG-1, KG-1a and SAML-2 AML cell lines with deletions of the long arm of one chromosome 5 (del(5q)) was performed. It has been shown that during long-term cultivation: 1) the cells retained diploid or near-diploid number of chromosomes; 2) unbalanced rearrangements of chromosomes 5, 7, 8, 11 and 17 were found in the karyotypes of all the cell lines examined; 3) involvement of intact chromosomes in novel chromosomal rearrangements was leukemia-associated; 4) chromosome imbalances restored AML-associated pattern; moreover, overrepresentation of distal parts of long arms of chromosomes 8 and 11 occurred.      The current data demonstrated the similar pattern of karyotypic variability, that drives the karyotypic progression of AML cell lines with del(5q) during long-term cultivation. Besides, it seems to be reasonable to use the panel of tumor cell lines with common primary cytogenetic abnormality for analysis of karyotypic variability of tumor cells in vitro. Key words:  acute myeloid leukemia, cell lines GF-D8, UoC-M1, HL-60, KG-1, KG-1a, SAML-2, deletions of chromosome 5q, karyotypic variability BIOLOGICAL PROPERTIES AND CRYOPRESERVATION OF PROTOZOAN PARASITE TRICHOMONAS VAGINALIS L.F. Litvinchuk1, N.V. Razdolskaya2, M.V. Potapchuk1, O.V. Gavrilova2, A.M. Ivanov2 1 Influenza Research Institute, 2 Military Medical Academy named after S.M. Kirov, St. Petersburg, Russia; office@influenza.spb.ru      Trichomonas vaginalis (Donne 1836) is a 7-30 µm sized flagellated protozoan, a human pathogen, causing 25 per cent of sexually transmitted infection of the urogenital tract.      The successful cryopreservation of T. vaginalis remains a significant problem of long-term preservation. In this study we examined the effect of freesing conditions and techniques, using for cell culture: cryoprotectant DMSO, medium for cell culture, regimen of cooling and thawing rates and finally deep freezing in liquid nitrogen.      We obtained six strains of trichomonads, freshly isolated from of patients with acute disease and incubated then at TYM (medium for isolation) for 3-4 days at 37 °C. A total of 5 x 104-105/ml parasites were added to the L-41 cells monolayers. Growth of the trichomonads population in L-41 cultures destroyed monolayers during 24-48 h. Passages of the protozoan were performed seweral times by transferring pathogen-containing suspension and cell debris of the previous L-41 culture into a new monolayer of L-41. This procedure has been effective for T. vaginalis. The final suspension for freesing contained cell culture medium (70-75%), active trichomonads (107-108/ml), L-41 cells, DMSO (5%), serum (20%). The recovery after cryopreservation for 2-3 years at -193 °C (liquid nitrogen) gave positive results and a high percent viability. The problems of long-term preservation in deep-frosen condition of T. vaginalis are discussed. Key words:  cultivation of trichomonas, сryoprotectant, сryopreservation, freezing, deep-frosen, Trichomonas banking STUDY OF THE BIOLOGICAL PROPERTIES OF RAT PROGENITOR NEURAL CELLS SUPERNATANT ON BRAIN GLIOMA CULTURES V.M. Semenova, L.D. Lyubych, N.I. Lisyany, A.Y. Glavatskiy, L.P. Stayno SI "Institute of Neurosurgery n.acad.A.P.Romodanov Acad. Med. Sci. of Ukraine", Kyiv, Ukraine; seveme22@rambler.ru      Purpose of the paper was the study of the rat progenitor neurocell (E12-16) supernatant effect on the growth activity of glial tumors in culture conditions. The dose-dependent effect of neurocell (E12-16) supernatant antitumour influence intensification was registered following 24-hour incubation with anaplastic glioma cultures. After incubation in presence with neurocell (E12-16) supernatant the dystrophic and necrobiotic changes in most tumors cells with disturbance of general structure of the cell growth area were observed intensifying under prolongation of incubation to 48 hours. Neurocell (E12-16) supernatant effect was dependent from the degree of tumor anaplasia. The most significant antitumor effect of the neurocell supernatant was registered in glioblastoma cultures. Key words:  rat progenitor neural cells, supernatant, biological properties, human brain glioma cultures, glioblastoma, antitumor effect SPECIFICS OF CHROMOSOMES' POSITION IN THE NUCLEUS OF HUMAN MESENCHIMAL STEM CELLS CULTIVATED IN VITRO A.V. Lavrov, Y.I. Voldgorn Research Center for Medical Genetics of RAMS, Moscow, Russia; avlavrov@yandex.ru      Modern studies focus on epigenetic mechanisms of gene regulation among which nuclear 3-D structure plays the most intriguing role. Adult stem cells are thought to be a very promising model for studying the influence of chromosome interactions on cell' properties including stem-cell properties and changes while diferentiation. In this work we studied position of chromosome centromers in the interphase nucleus while in vitro cultivating mesenchimal stem cells (MSC). More than 400 nuclei were analyzed in 9 cultures of MSC. Radial distances of centromers of chromosomes 6 were bigger (0,68) than of chromosome 18 (0,49). Homologues of each chromosome had different radial distances. Fat tissue and bone marrow derived MSC demonstrate no differences in radial distances of the studied chromosomes. However centromer 6 moved to periphery after cultivating for more than 8 passages, which is probably connected with age-related changes. Key words:  chromosome territory, nuclear structure, mesenchymal stem cell CELL LINE STANDARDIZATION OPPORTUNITIES BY GENOTYPING WITH SHORT TANDEMIC REPEATS A.Yu. Markaryan, A.A. Bakharev, P.V. Ustjanzev FSSI "Ekaterinburg Research Institute of Viral Infections" of Rospotrebnadzor, Ekaterinburg, Russia; virus@etel.ru      Results obtained by analysis of cell lines of human origin stored at the ERIVI bank-museum are presented. A polymerase chain reaction (PCR) technique employing monoplexes of human tetramer STR loci was used.      Primary analysis demonstrated absence of cross-contamination in the bank-museum cell lines investigated and detected genetic differences in some clones.      The research led us to establish partial coincidence in the lengths of amplified human STR loci fragments and African green monkey cell line (Vero). Presence of homologous genome fragments was suggested. Key words:  cell lines, PCR, STR loci DETECTION OF MYCOPLASMA AND BOVINE VIRAL DIARRHEA VIRUS CONTAMINATION IN THE COLLECTION OF CELL LINES R.Ya. Podchernyaeva, L.V. Uryvaev, A.V. Dedova, L.V. Dedova, K.S. Ionova, G.R. Michailova, N.A. Parasyuk, O.M. Grinkevich, T.K. Selivanova, T.V. Grebenikova, A.D. Petrachev, M.N. Shetvin The D.I. Ivanovsky Institute of Virology of the Ministry of Health and Social Development of the Russian Federation, Moscow, Russia; cells@rambler.ru       The paper contains the data for detection of the toxicity of fetal calf sera obtained from different firms. Micoplasma contamination of tissue cultures was defected by olivomycin and PCR methods. Contamination of 83 cellular lines by the bovine diarrhea virus was studied using PCR. It was shown that 30% of lines contain the virus. Key words:  cell cultures, toxicity of bovine sera, contamination, mycoplasma, bovine diarrhea virus INSTRUCTIONS TO AUTHORS      The paper should not exceed 10 manuscript pages. The manuscript is to be accompanied by recommendation addressed to the Editorial Board of the Information Bulletin from the institution in which the work has been carried out. The paper should contain a brief resume, with the tittle of the work, names of the authors, institution and city.      The manuscript is to be accompanied by its electron variant: one file in MS Word for Windows. The file should have the font Arial Narrow, 13; interline difference - 1.5; the indented line - 0.5 cm; the upper margin - 2.5 cm, other margins - 2.1 cm each. The file may be e-mailed to the address of the Editorial Board. Address of the Editorial Board: Dr. M. Bogdanova, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky Prosp. 4, St. Petersburg 194064, Russia. Tel.: (812) 297-53-10, 8-911-284-28-64, fax: (812) 297-03-41 e-mail: msb2051@rambler.ru