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Pathological changes in mice treated with cyclophosphamide and exogenous DNA

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Russian Journal of Genetics: Applied Research

Abstract

The synergistic action of the cytostatic drug cyclophosphamide (CP) and fragmented exogenous DNA causes illness and death in mice (Dolgova et al., 2011–2013). The observed “delayed death” effect was most clearly pronounced when the DNA preparation was administered 18 to 30 h after CP treatment. This time span was termed the “death window.”

It was found that injections of exogenous DNA result in a sustained increase in bone-marrow cell (BMC) apoptosis, which occurs throughout the time of DNA administration (18–30 h). Exogenous DNA, both allogeneic and belonging to various taxa, induces BMC apoptosis. Plasmid DNA has the greatest effect on apoptosis induction. The analysis of reduction and restoration of the BMC subpopulations as the mice progressed to death revealed a virtually complete loss of the 12–20-μm fraction of the cell population (about 3–4% vs. 35–40% in the control), which corresponds to the maximum leukopenia on day 3 after CP treatment. However, the relative number of CD34+ hematopoietic stem cells (HSCs) from day 15 and until the end of the observation constituted 1.2–1.4%, which corresponds to the wild-type range. Comparison of BMC smears from the sternal bone marrow of the CP and CP + DNA groups of mice indicated that the BMC populations isolated from CP + DNA animals lack young committed lymphopoiesis progenitor cells. Moreover, the affected mice had immature blast cell types in their blood, which was never observed in healthy or CP-treated mice. Pathological and morphological analyses show that starting from posttreatment day 9, mice that received CP + DNA preparations displayed pronounced morphological changes in their lungs, liver, pancreas, central and peripheral immune system organs, and brain. Most of the pathological changes observed are consistent with a severe inflammatory response. This suggestion was proven by structural equivalents of functional involution of lymphoid organs, such as the thymus, spleen, and lymph nodes.

We speculate that the death of treated animals resulted from multiple organ dysfunctions caused by accidental involution of lymphoid organs and the systemic inflammatory response syndrome, both associated with injections of fragmented exogenous DNA into experimental animals within the “death window,” which corresponds to the final step in the repair of the majority of CP-induced double-strand breaks.

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References

  • Afanas’eva, Yu.I., Kuznetsova, S.L., and Yurina, N.A., Gistologiya, tsitologiya i embriologiya (Histology, Cytology, and Embryology), Moscow: Meditsina, 2004.

    Google Scholar 

  • Andrews, N.W., Membrane repair and immunological danger, EMBO Rep., 2005, vol. 6, no. 9, pp. 826–830.

    Article  PubMed  CAS  Google Scholar 

  • Buckley, R.H., Primary immunodeficiency diseases due to defects in lymphocytes, N. Engl. J. Med., 2000, vol. 343, no. 18, pp. 1313–1324.

    Article  PubMed  CAS  Google Scholar 

  • Coban, C., Koyama, S., Takeshita, F., et al., Molecular and cellular mechanisms of DNA vaccines, Hum. Vaccin., 2008, vol. 4, pp. 453–456.

    Article  PubMed  CAS  Google Scholar 

  • Cook, R., Wu, C.C., Kang, Y.J., and Han, J., The role of the p38 pathway in adaptive immunity, Cell. Mol. Immunol., 2007, vol. 4, no. 4, pp. 253–259.

    PubMed  CAS  Google Scholar 

  • Decker, P., Wolburg, H., and Rammensee, H.G., Nucleosomes induce lymphocyte necrosis, Eur. J. Immunol., 2003, vol. 33, no. 7, pp. 1978–1987.

    Article  PubMed  Google Scholar 

  • Decker, P., Singh-Jasuja, H., Haager, S., et al., Nucleosome, the main autoantigen in systemic lupus erythematosis, induces direct dendritic cell activation via a MyD88-independent pathway: consequences on inflammation, J. Immunol., 2005, vol. 174, no. 6, pp. 3326–3334.

    PubMed  CAS  Google Scholar 

  • Derbyshire, M.K., Epstein, L.H., Young, C.S.H., et al., Nonhomologous recombination in human cells, Mol. Cell Biol., 1994, vol. 14, no. 1, pp. 156–169.

    PubMed  CAS  Google Scholar 

  • Dolgova, E.V., Prokopenko, A.V., Nikolin, V.P., et al., Characterization of changes in the number of moderate repeats in the genome of bone marrow cells of experimental mice treated with cyclophosphamide and exogenous human DNA, Russ. J. Genet. Appl. Res., 2013 (in press).

    Google Scholar 

  • Dolgova, E.V., Proskurina, A.S., Nikolin, V.P., et al., Characterization of temporal parameters of manifestations of the toxic effect of injections of exogenous DNA after pretreatment with the cytostatic cyclophosphamide, Vavilov. Zh. Genet. Selekts., 2011, vol. 15, no. 4, pp. 674–689.

    Google Scholar 

  • Dolgova, E.V., Nikolin, V.P., Popova, N.A., et al., Internalization of exogenous DNA into internal compartments of murine bone marrow cells, Russ. J. Genet. Appl. Res., 2012, vol. 2, no. 6, pp. 440–452.

    Article  Google Scholar 

  • Fugmann, S.D., Rag1 and rag2 in V (D) J recombination and transposition, Immunol. Res., 2001, vol. 23, no. 1, pp. 23–39.

    Article  PubMed  CAS  Google Scholar 

  • De Gregorio, E. and Rappuoli, R., Inside sensors detecting outside pathogens, Nat. Immunol., 2004, vol. 5, no. 11, pp. 1099–1100.

    Article  PubMed  Google Scholar 

  • Harty, J.T., Tvinnereim, A.R., and White, D.W., CD8+ T cell effector mechanisms in resistance to infection, Annu. Rev. Immunol., 2000, vol. 18, pp. 275–308.

    Article  PubMed  CAS  Google Scholar 

  • Ishii, K.J. and Akira, S., Innate immune recognition of, and regulation by, DNA, Trends Immunol., 2006, vol. 27, pp. 525–532.

    Article  PubMed  CAS  Google Scholar 

  • Kaufmann, S.H. and Schaible, U.E., Antigen presentation and recognition in bacterial infections, Curr. Opin. Immunol., 2005, vol. 17, no. 1, pp. 79–87.

    Article  PubMed  CAS  Google Scholar 

  • Kotnis, A., Du, L., Liu, C., et al., Non-homologous end joining in class switch recombination: the beginning of the end, Philos. Trans. R. Soc. Lond. B. Biol. Sci., 2009, vol. 364, no. 1517, pp. 653–665.

    Article  PubMed  CAS  Google Scholar 

  • Krishan, A., Rapid flow cytofluorometric analysis of mammalian cell cycle by propidium iodide staining, J. Cell Biol., 1975, vol. 66, no. 1, pp. 188–193.

    Article  PubMed  CAS  Google Scholar 

  • Kruglyakov, P.V., Sokolova, I.B., and Polyntsev, D.G., Stem cells of differentiating tissues of an adult organism, Tsitologiya, 2008, vol. 50, no. 7, pp. 557–567.

    Google Scholar 

  • Lee, S., Oshige, M., Durant, S.T., et al., The SET domain protein Metnase mediates foreign DNA integration and links integration to nonhomologous end-joining repair, Proc. Natl. Acad. Sci. U.S.A., 2005, vol. 102, no. 50, pp. 18075–18080.

    Article  PubMed  CAS  Google Scholar 

  • Lees-Miller, S.P. and Meek, K., Repair of DNA double strand breaks by non-homologous end joining, Biochimie, 2003, vol. 85, no. 11, pp. 1161–1173.

    Article  PubMed  CAS  Google Scholar 

  • Lekstrom-Himes, J.A. and Gallin, J.I., Immunodeficiency diseases caused by defects in phagocytes, N. Engl. J. Med., 2000, vol. 343, no. 23, pp. 1703–1714.

    Article  PubMed  CAS  Google Scholar 

  • Macgregor, C. and Varley, J., Working with Animal Chromosomes, Chichester: Wiley, 1984.

    Google Scholar 

  • Martin, D.A. and Elkon, K.B., Intracellular mammalian DNA stimulates myeloid dendritic cells to produce type I interferons predominantly through a toll-like receptor 9-independent pathway, Arthritis Rheum., 2006, vol. 54, pp. 951–962.

    Article  PubMed  CAS  Google Scholar 

  • Medzhitov, R., Recognition of microorganisms and activation of the immune response, Nature, 2007, vol. 449, no. 7164, pp. 819–826.

    Article  PubMed  CAS  Google Scholar 

  • Mayle, D., Brostoff, J., Roth, D.B., and Reutte, A., Immunologiya (Immunology), Moscow: Logosfera, 2007.

    Google Scholar 

  • Merrell, D.S. and Falkow, S., Frontal and stealth attack strategies in microbial pathogenesis, Nature, 2004, vol. 430, no. 6996, pp. 250–256.

    Article  PubMed  CAS  Google Scholar 

  • Mueller, A. and Falkow, S., Persistent bacterial infections: the interface of the pathogen and the host immune system, Nat. Rev. Microbiol., 2004, vol. 2, no. 9, pp. 747–765.

    Article  PubMed  Google Scholar 

  • Napirei, M., Karsunky, H., Zevnik, B., et al., Features of systemic lupus erythematosis in DNase 1-deficient mice, Nat. Genet., 2000, vol. 25, no. 2, pp. 177–181.

    Article  PubMed  CAS  Google Scholar 

  • Orkin, S.H. and Zon, L.I., Hematopoiesis: an evolving paradigm for stem cell biology, Cell, 2008, vol. 132, no. 4, pp. 631–644.

    Article  PubMed  CAS  Google Scholar 

  • Proskuryakov, S.Ya., Gavai, V.P., and Konoplyannikov, A.G., Immunology of necrosis and apoptosis, Biochemistry (Moscow), 2005, vol. 70, pp. 1593–1605.

    Google Scholar 

  • Ravetch, J.V., A full complement of receptors in immune complex diseases, J. Clin. Invest., 2002, vol. 110, no. 12, pp. 1759–1761.

    PubMed  CAS  Google Scholar 

  • Rosen, F.S., Cooper, M.D., and Wedgwood, R.J., The primary immunodeficiencies, N. Engl. J. Med., 1995, vol. 333, no. 7, pp. 431–440.

    Article  PubMed  CAS  Google Scholar 

  • Rossi, D.J., Jamieson, C.H., and Weissman, I.L., Stems cells and the pathways to aging and cancer, Cell, 2008, vol. 132, no. 4, pp. 681–696.

    Article  PubMed  CAS  Google Scholar 

  • Rouse, B.T. and Suvas, S., Regulatory cells and infectious agents: detentes cordiale and contraire, J. Immunol., 2004, vol. 173, no. 4, pp. 2211–2215.

    PubMed  CAS  Google Scholar 

  • Rykova, E.Yu., Laktionov, P.P., and Vlasov, V.V., The activatory effect of DNA on the immune system, Usp. Sovrem. Biol., 2001, vol. 121, pp. 160–171.

    CAS  Google Scholar 

  • Sambrook, J., Fritsch, E.F., and Maniatis, T., Molecular Cloning, a Laboratory Manual, 2nd ed., Cold Spring Harbor: Cold Spring Harbor Laboratory Press, 1989.

    Google Scholar 

  • Shirota, H., Ishii, K.J., Takakuwa, H., and Klinman, D.M., Contribution of interferon-beta to the immune activation induced by double-stranded DNA, Immunology, 2006, vol. 118, pp. 302–310.

    Article  PubMed  CAS  Google Scholar 

  • Silva, J. and Smith, A., Capturing pluripotency, Cell, 2008, vol. 132, no. 4, pp. 532–536.

    Article  PubMed  CAS  Google Scholar 

  • Takeshita, F. and Ishii, K.J., Intracellular DNA sensors in immunity, Curr. Opin. Immunol., 2008, vol. 20, pp. 383–388.

    Article  PubMed  CAS  Google Scholar 

  • Volkova, O.V. and Eletskii, Yu.K., Osnovy gistologii i gistologicheskoi tekhniki (Fundamentals of Histology and Histological Techniques), Moscow: Meditsina, 1971.

    Google Scholar 

  • Wang, H., Rosidi, B., Perrault, R., et al., DNA ligase III as a candidate component of backup pathways of nonhomologous end joining, Cancer Res., 2005, vol. 65, no. 10, pp. 4020–4030.

    Article  PubMed  CAS  Google Scholar 

  • Warren, J.S., Yabroff, K.R., Remickd, G., et al., Tumor necrosis factor participates in the pathogenesis of acute immune complex alveolitis in the rat, J. Clin. Invest., 1989, vol. 84, no. 6, pp. 1873–1882.

    Article  PubMed  CAS  Google Scholar 

  • Whaley, K., Complement and immune complex diseases, in Complement in Health and Disease, Whaley, K., Ed., Lancaster: MTP Press Ltd, 1987.

    Google Scholar 

  • Yu, J. and Thomson, J.A., Pluripotent stem cell lines, Genes Dev., 2008, vol. 22, no. 15, pp. 1987–1997.

    Article  PubMed  CAS  Google Scholar 

Download references

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Correspondence to E. V. Dolgova.

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Original Russian Text © E.V. Dolgova, V.P. Nikolin, N.A. Popova, A.S. Proskurina, K.E. Orishchenko, E.A. Alyamkina, Ya.R. Efremov, S.I. Baiborodin, E.R. Chernykh, A.A. Ostanin, S.S. Bogachev, T.S. Gvozdeva, E.M. Malkova, O.S. Taranov, V.A. Rogachev, A.S. Panov, S.N. Zagrebelnyi, M.A. Shurdov, 2013, published in Vavilovskii Zhurnal Genetiki i Selektsii, 2013, Vol. 17, No. 1, pp. 129–146.

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Dolgova, E.V., Nikolin, V.P., Popova, N.A. et al. Pathological changes in mice treated with cyclophosphamide and exogenous DNA. Russ J Genet Appl Res 3, 291–304 (2013). https://doi.org/10.1134/S2079059713040035

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  • DOI: https://doi.org/10.1134/S2079059713040035

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