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Biochemistry (Moscow)

, Volume 84, Issue 3, pp 263–271 | Cite as

Alteration of Hypoxia-Associated Gene Expression in Replicatively Senescent Mesenchymal Stromal Cells under Physiological Oxygen Level

  • A. Yu. Ratushnyy
  • Yu. V. Rudimova
  • L. B. BuravkovaEmail author
Article
  • 11 Downloads

Abstract

Mesenchymal stromal cells (MSCs) are a population of adult stem cells that modulate functional state of neighboring tissues. During cell aging, the biological activity of MSC changes, which may affect tissue homeostasis. It is known that reducing the oxygen level in vitro to physiological values typical to a particular cell niche leads to attenuation of some morphological and functional changes associated with aging. This work aimed to study gene expression in MSCs involved in response to physiological hypoxia using a replicative aging model under physiological (5%) and atmospheric (20%) oxygen in cultures. Our results show that significant reduction of proliferative activity of MSCs is observed after 20 passages (~50 cell generations). Regardless of the oxygen, in senescent cells PKM2, SERPINE1, and VEGFA were upregulated while ANKRD37, DDIT4, HIF1A, and TXNIP were downregulated. Also, ADORA2B, BNIPL, CCNG2, EGLN1, MAP3K1, MXI1, and P4HA1 were downregulated under hypoxia. The effect of oxygen was more pronounced at earlier passages both on the cellular and transcription levels. Irrespective of the passage, genes ANGPTL4, GYS1, PKM2, SERPINE1, and TP53 were downregulated under hypoxia. Also, decreased expression was observed for ADM, F10, HMOX1, P4HB, PFKL, SLC16A3 in earlier passages, and for HK2 – in later passages. Upregulation was only observed for ANKRD37, both at early and late cultures.

Keywords

mesenchymal stromal cells replicative aging oxygen level 

Abbreviations

HIF

hypoxia-induced factor

MSCs

mesenchy-mal stromal cells

PD

population doubling

SA-β-gal

senescence-associated β-galactosidase

SASP

senescence-associated secretory phenotype

senMSCs

senescent MSCs

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References

  1. 1.
    Munoz–Espin, D., and Serrano, M. (2014) Cellular senes–cence: from physiology to pathology, Nat. Rev. Mol. Cell Biol., 15, 482–496.CrossRefGoogle Scholar
  2. 2.
    Lopez–Otin, C., Blasco, M. A., Partridge, L., Serrano, M., and Kroemer, G. (2013) The hallmarks of aging, Cell, 153, 1194–1217.CrossRefGoogle Scholar
  3. 3.
    McHugh, D., and Gil, J. (2018) Senescence and aging: causes, consequences, and therapeutic avenues, J. Cell Biol., 217, 65–77.Google Scholar
  4. 4.
    Campisi, J., and d’Adda di Fagagna, F. (2007) Cellular senescence: when bad things happen to good cells, Nat. Rev. Mol. Cell Biol., 8, 729–740.CrossRefGoogle Scholar
  5. 5.
    Collado, M., Blasco, M. A., and Serrano, M. (2007) Cellular senescence in cancer and aging, Cell, 130, 223–233.CrossRefGoogle Scholar
  6. 6.
    Salama, R., Sadaie, M., Hoare, M., and Narita, M. (2014) Cellular senescence and its effector programs, Genes Dev., 28, 99–114.CrossRefGoogle Scholar
  7. 7.
    Watanabe, S., Kawamoto, S., Ohtani, N., and Hara, E. (2017) Impact of senescence–associated secretory pheno–type and its potential as a therapeutic target for senescence–associated diseases, Cancer Sci., 108, 563–569.CrossRefGoogle Scholar
  8. 8.
    Kuilman, T., and Peeper, D. S. (2009) Senescence–messag–ing secretome: SMS–ing cellular stress, Nat. Rev. Cancer, 9, 81–94.CrossRefGoogle Scholar
  9. 9.
    Coppe, J. P., Desprez, P. Y., Krtolica, A., and Campisi, J. (2010) The senescence–associated secretory phenotype: the dark side of tumor suppression, Annu. Rev. Pathol., 5, 99–118.CrossRefGoogle Scholar
  10. 10.
    Minieri, V., Saviozzi, S., Gambarotta, G., Lo Iacono, M., Accomasso, L., Cibrario Rocchietti, E., Gallina, C., Turinetto, V., and Giachino, C. (2015) Persistent DNA damage–induced premature senescence alters the function–al features of human bone marrow mesenchymal stem cells, J. Cell Mol. Med., 19, 734–743.CrossRefGoogle Scholar
  11. 11.
    Turinetto, V., Vitale, E., and Giachino, C. (2016) Senescence in human mesenchymal stem cells: functional changes and implications in stem cell–based therapy, Int. J. Mol. Sci., 17, E1164.Google Scholar
  12. 12.
    Wei, F., Qu, C., Song, T., Ding, G., Fan, Z., Liu, D., Liu, Y., Zhang, C., Shi, S., and Wang, S. (2012) Vitamin C treatment promotes mesenchymal stem cell sheet forma–tion and tissue regeneration by elevating telomerase activi–ty, J. Cell Physiol., 227, 3216–3224.CrossRefGoogle Scholar
  13. 13.
    Lin, T. M., Tsai, J. L., Lin, S. D., Lai, C. S., and Chang, C. C. (2005) Accelerated growth and prolonged lifespan of adipose tissue–derived human mesenchymal stem cells in a medium using reduced calcium and antioxidants, Stem Cells Dev., 14, 92–102.CrossRefGoogle Scholar
  14. 14.
    Skulachev, V. P. (2013) Cationic antioxidants as a powerful tool against mitochondrial oxidative stress, Biochem. Biophys. Res. Commun., 441, 275–279.CrossRefGoogle Scholar
  15. 15.
    Skulachev, M. V., and Skulachev, V. P. (2017) Programmed aging of mammals: proof of concept and prospects of bio–chemical approaches for anti–aging therapy, Biochemistry (Moscow), 82, 1403–1422.CrossRefGoogle Scholar
  16. 16.
    Fehrer, C., Brunauer, R., Laschober, G., Unterluggauer, H., Reitinger, S., Kloss, F., Gully, C., Gassner, R., and Lepperdinger, G. (2007) Reduced oxygen tension attenu–ates differentiation capacity of human mesenchymal stem cells and prolongs their lifespan, Aging Cell., 6, 745–757.CrossRefGoogle Scholar
  17. 17.
    Choi, J. R., Pingguan–Murphy, B., Wan Abas, W. A., Yong, K. W., Poon, C. T., Noor Azmi, M. A., Omar, S. Z., Chua, K. H., Xu, F., and Wan Safwani, W. K. (2015) In situ nor–moxia enhances survival and proliferation rate of human adipose tissue–derived stromal cells without increasing the risk of tumorigenesis, PLoS One, 10, e0115034.Google Scholar
  18. 18.
    Buravkova, L. B., Andreeva, E. R., Gogvadze, V., and Zhivotovsky, B. (2014) Mesenchymal stem cells and hypox–ia: where are we? Mitochondrion, 19, 105–112.CrossRefGoogle Scholar
  19. 19.
    Lobanova, M. V., Ratushnyy, A. Y., and Buravkova, L. B. (2016) Expression of senescence–associated genes in multi–potent mesenchymal stromal cells during long–term culti–vation at various hypoxic levels, Dokl. Biochem. Biophys., 470, 326–328.CrossRefGoogle Scholar
  20. 20.
    Ratushnyy, A., Lobanova, M., and Buravkova, L. B. (2017) Expansion of adipose tissue–derived stromal cells at “phys–iologic” hypoxia attenuates replicative senescence, Cell Biochem. Funct., 35, 232–243.CrossRefGoogle Scholar
  21. 21.
    Semenza, G. L. (2007) Hypoxia–inducible factor 1 (HIF–1) pathway, Sci. STKE, 2007, cm8.CrossRefGoogle Scholar
  22. 22.
    Zuk, P. A., Zhu, M., Mizuno, H., Huang, J., Futrell, J. W., Katz, A. J., Benhaim, P., Lorenz, H. P., and Hedrick, M. H. (2001) Multilineage cells from human adipose tissue: implications for cell–based therapies, Tissue Eng., 7, 211–228.CrossRefGoogle Scholar
  23. 23.
    Buravkova, L. B., Grinakovskaya, O. S., Andreeva, E. R., Zhambalova, A. P., and Kozionova, M. P. (2009) Characteristics of human lipoaspirate–isolated mesenchy–mal stromal cells cultivated under lower oxygen tension, Tsitologiya, 51, 4–10.Google Scholar
  24. 24.
    Dominici, M., Le Blanc, K., Mueller, I., Slaper–Cortenbach, I., Marini, F., Krause, D., Deans, R., Keating, A., Prockop, D. J., and Horwitz, E. (2006) Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement, Cytotherapy, 8, 315–317.CrossRefGoogle Scholar
  25. 25.
    Greenwood, S. K., Hill, R. B., Sun, J. T., Armstrong, M. J., Johnson, T. E., Gara, J. P., and Galloway, S. M. (2004) Population doubling: a simple and more accurate estima–tion of cell growth suppression in the in vitro assay for chro–mosomal aberrations that reduces irrelevant positive results, Environ. Mol. Mutagen., 43, 36–44.CrossRefGoogle Scholar
  26. 26.
    Livak, K. J., and Schmittgen, T. D. (2001) Analysis of rela–tive gene expression data using real–time quantitative PCR and the 2–ΔΔCT method, Methods, 25, 402–408.CrossRefGoogle Scholar
  27. 27.
    McLeod, C. M., and Mauck, R. L. (2017) On the origin and impact of mesenchymal stem cell heterogeneity: new insights and emerging tools for single cell analysis, Eur. Cell Mater., 34, 217–231.CrossRefGoogle Scholar
  28. 28.
    Gonzalez–Cruz, R. D., Fonseca, V. C., and Darling, E. M. (2012) Cellular mechanical properties reflect the differenti–ation potential of adipose–derived mesenchymal stem cells, Proc. Natl. Acad. Sci. USA, 109, E1523–E1529.Google Scholar
  29. 29.
    Dimri, G. P., Lee, X., Basile, G., Acosta, M., Scott, G., Roskelley, C., Medrano, E. E., Linskens, M., Rubelj, I., Pereira–Smith, O., Peacocke, M., and Campisi, J. (1995) A biomarker that identifies senescent human cells in culture and in aging skin in vivo, Proc. Natl. Acad. Sci. USA, 92, 9363–9367.CrossRefGoogle Scholar
  30. 30.
    Xu, L. N., Lin, N., Xu, B. N., Li, J. B., and Chen, S. Q. (2015) Effect of human umbilical cord mesenchymal stem cells on endometriotic cell proliferation and apoptosis, Genet. Mol. Res., 14, 16553–16561.CrossRefGoogle Scholar
  31. 31.
    Bakkenist, C. J., and Kastan, M. B. (2004) Phosphatases join kinases in DNA–damage response pathways, Trends Cell Biol., 14, 339–341.CrossRefGoogle Scholar
  32. 32.
    Zhan, H., Suzuki, T., Aizawa, K., Miyagawa, K., and Nagai, R. (2010) Ataxia telangiectasia mutated (ATM)–mediated DNA damage response in oxidative stress–induced vascular endothelial cell senescence, J. Biol. Chem., 285, 29662–29670.CrossRefGoogle Scholar
  33. 33.
    Buscemi, G., Perego, P., Carenini, N., Nakanishi, M., Chessa, L., Chen, J., Khanna, K., and Delia, D. (2004) Activation of ATM and Chk2 kinases in relation to the amount of DNA strand breaks, Oncogene, 23, 7691–7700.CrossRefGoogle Scholar
  34. 34.
    Lukas, C., Falck, J., Bartkova, J., Bartek, J., and Lukas, J. (2003) Distinct spatiotemporal dynamics of mammalian checkpoint regulators induced by DNA damage, Nat. Cell Biol., 5, 255–260.CrossRefGoogle Scholar
  35. 35.
    Von Zglinicki, T., Saretzki, G., Ladhoff, J., d’Adda di Fagagna, F., and Jackson, S. P. (2005) Human cell senes–cence as a DNA damage response, Mech. Ageing Dev., 126, 111–117.CrossRefGoogle Scholar
  36. 36.
    Sofer, A., Lei, K., Johannessen, C. M., and Ellisen, L. W. (2005) Regulation of mTOR and cell growth in response to energy stress by REDD1, Mol. Cell Biol., 25, 5834–5845.CrossRefGoogle Scholar
  37. 37.
    Shoshani, T., Faerman, A., Mett, I., Zelin, E., Tenne, T., Gorodin, S., Moshel, Y., Elbaz, S., Budanov, A., Chajut, A., Kalinski, H., Kamer, I., Rozen, A., Mor, O., Keshet, E., Leshkowitz, D., Einat, P., Skaliter, R., and Feinstein, E. (2002) Identification of a novel hypoxia–inducible factor 1–responsive gene, RTP801, involved in apoptosis, Mol. Cell Biol., 22, 2283–2293.CrossRefGoogle Scholar
  38. 38.
    Wolff, N. C., Vega–Rubin–de–Celis, S., Xie, X. J., Castrillon, D. H., Kabbani, W., and Brugarolas, J. (2011) Cell–type–dependent regulation of mTORC1 by REDD1 and the tumor suppressors TSC1/TSC2 and LKB1 in response to hypoxia, J. Mol. Cell Biol., 31, 1870–1884.CrossRefGoogle Scholar
  39. 39.
    Kucejova, B., Sunny, N. E., Nguyen, A. D., Hallac, R., Fu, X., Pena–Llopis, S., Mason, R. P., Deberardinis, R. J., Xie, X. J., Debose–Boyd, R., Kodibagkar, V. D., Burgess, S. C., and Brugarolas, J. (2011) Uncoupling hypoxia signaling from oxygen sensing in the liver results in hypoketotic hypoglycemic death, Oncogene, 30, 2147–2160.CrossRefGoogle Scholar
  40. 40.
    Benita, Y., Kikuchi, H., Smith, A. D., Zhang, M. Q., Chung, D. C., and Xavier, R. J. (2009) An integrative genomics approach identifies hypoxia inducible factor–1 (HIF–1)–target genes that form the core response to hypox–ia, Nucleic Acids Res., 37, 4587–4602.CrossRefGoogle Scholar
  41. 41.
    Murakami, S., Terakura, M., Kamatani, T., Hashikawa, T., Saho, T., Shimabukuro, Y., and Okada, H. (2000) Adenosine regulates the production of interleukin–6 by human gingival fibroblasts via cyclic AMP/protein kinase A pathway, J. Periodont. Res., 35, 93–101.CrossRefGoogle Scholar
  42. 42.
    Ray, R., Chen, G., Vande Velde, C., Cizeau, J., Park, J. H., Reed, J. C., Gietz, R. D., and Greenberg, A. H. (2000) BNIP3 heterodimerizes with Bcl–2/Bcl–X(L) and induces cell death independent of a Bcl–2 homology 3 (BH3) domain at both mitochondrial and nonmitochondrial sites, J. Biol. Chem., 275, 1439–1448.CrossRefGoogle Scholar
  43. 43.
    Ahmed, S., Al–Saigh, S., and Matthews, J. (2012) FOXA1 is essential for aryl hydrocarbon receptor–dependent regu–lation of cyclin G2, Mol. Cancer Res., 10, 636–648.CrossRefGoogle Scholar
  44. 44.
    Sun, G. G., Zhang, J., and Hu, W. N. (2014) CCNG2 expression is downregulated in colorectal carcinoma and its clinical significance, Tumour Biol., 35, 3339–3346.CrossRefGoogle Scholar
  45. 45.
    Huang, H. Y., Chen, S. Z., Zhang, W. T., Wang, S. S., Liu, Y., Li, X., Sun, X., Li, Y. M., Wen, B., Lei, Q. Y., and Tang, Q. Q. (2013) Induction of EMT–like response by BMP4 via up–regulation of lysyl oxidase is required for adipocyte lin–eage commitment, Stem Cell Res., 10, 278–287.CrossRefGoogle Scholar
  46. 46.
    Kortlever, R. M., Higgins, P. J., and Bernards, R. (2006) Plasminogen activator inhibitor–1 is a critical downstream target of p53 in the induction of replicative senescence, Nat. Cell Biol., 8, 877–884.CrossRefGoogle Scholar
  47. 47.
    Zhang, Y., Xu, Y., Ma, J., Pang, X., and Dong, M. (2017) Adrenomedullin promotes angiogenesis in epithelial ovari–an cancer through upregulating hypoxia–inducible factor–1α and vascular endothelial growth factor, Sci. Rep., 7, 40524.CrossRefGoogle Scholar
  48. 48.
    Chen, K., and Maines, M. D. (2000) Nitric oxide induces heme oxygenase–1 via mitogen–activated protein kinases ERK and p38, Cell Mol. Biol. (Noisy–le–grand), 46, 609–617.Google Scholar
  49. 49.
    Pescador, N., Villar, D., Cifuentes, D., Garcia–Rocha, M., Ortiz–Barahona, A., Vazquez, S., Ordonez, A., Cuevas, Y., Saez–Morales, D., Garcia–Bermejo, M. L., Landazuri, M. O., Guinovart, J., and del Peso, L. (2010) Hypoxia pro–motes glycogen accumulation through hypoxia inducible factor (HIF)–mediated induction of glycogen synthase 1, PLoS One, 5, e9644.Google Scholar
  50. 50.
    Kim, I., Kim, H. G., Kim, H., Kim, H. H., Park, S. K., Uhm, C. S., Lee, Z. H., and Koh, G. Y. (2000) Hepatic expression, synthesis and secretion of a novel fibrinogen/angiopoietin–related protein that prevents endothelial cell apoptosis, Biochem. J., 346, 603–610.CrossRefGoogle Scholar
  51. 51.
    Pogodina, M. V., and Buravkova, L. B. (2015) Expression of HIF–1α in multipotent mesenchymal stromal cells under hypoxic conditions, Bull. Exp. Biol. Med., 159, 355–357.CrossRefGoogle Scholar
  52. 52.
    Tsai, C. C., Chen, Y. J., Yew, T. L., Chen, L. L., Wang, J. Y., Chiu, C. H., and Hung, S. C. (2011) Hypoxia inhibits senescence and maintains mesenchymal stem cell properties through down–regulation of E2A–p21 by HIF–TWIST, Blood, 117, 459–469.CrossRefGoogle Scholar

Copyright information

© Pleiades Publishing, Ltd. 2019

Authors and Affiliations

  • A. Yu. Ratushnyy
    • 1
  • Yu. V. Rudimova
    • 1
  • L. B. Buravkova
    • 1
    Email author
  1. 1.Institute of Biomedical ProblemsRussian Academy of SciencesMoscowRussia

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