Skip to main content
SpringerLink
Log in

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

  • Published:
Biochemistry (Moscow) Aims and scope Submit manuscript

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.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

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

References

  1. Munoz–Espin, D., and Serrano, M. (2014) Cellular senes–cence: from physiology to pathology, Nat. Rev. Mol. Cell Biol., 15, 482–496.

    Article  CAS  PubMed  Google Scholar 

  2. Lopez–Otin, C., Blasco, M. A., Partridge, L., Serrano, M., and Kroemer, G. (2013) The hallmarks of aging, Cell, 153, 1194–1217.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. McHugh, D., and Gil, J. (2018) Senescence and aging: causes, consequences, and therapeutic avenues, J. Cell Biol., 217, 65–77.

    CAS  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  5. Collado, M., Blasco, M. A., and Serrano, M. (2007) Cellular senescence in cancer and aging, Cell, 130, 223–233.

    Article  CAS  PubMed  Google Scholar 

  6. Salama, R., Sadaie, M., Hoare, M., and Narita, M. (2014) Cellular senescence and its effector programs, Genes Dev., 28, 99–114.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Kuilman, T., and Peeper, D. S. (2009) Senescence–messag–ing secretome: SMS–ing cellular stress, Nat. Rev. Cancer, 9, 81–94.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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. 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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  14. Skulachev, V. P. (2013) Cationic antioxidants as a powerful tool against mitochondrial oxidative stress, Biochem. Biophys. Res. Commun., 441, 275–279.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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. 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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  21. Semenza, G. L. (2007) Hypoxia–inducible factor 1 (HIF–1) pathway, Sci. STKE, 2007, cm8.

    Book  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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. 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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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. 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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  31. Bakkenist, C. J., and Kastan, M. B. (2004) Phosphatases join kinases in DNA–damage response pathways, Trends Cell Biol., 14, 339–341.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  Google Scholar 

  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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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.

    Article  CAS  Google Scholar 

  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.

    Article  CAS  Google Scholar 

  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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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.

    Article  CAS  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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.

    CAS  Google Scholar 

  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. 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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

  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.

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Institute of Biomedical Problems, Russian Academy of Sciences, 123007, Moscow, Russia

    A. Yu. Ratushnyy, Yu. V. Rudimova & L. B. Buravkova

Authors
  1. A. Yu. Ratushnyy
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yu. V. Rudimova
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. L. B. Buravkova
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to L. B. Buravkova.

Additional information

Russian Text © A. Yu. Ratushnyy, Yu. V. Rudimova, L. B. Buravkova, 2019, published in Biokhimiya, 2019, Vol. 84, No. 3, pp. 380–391.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Ratushnyy, A.Y., Rudimova, Y.V. & Buravkova, L.B. Alteration of Hypoxia-Associated Gene Expression in Replicatively Senescent Mesenchymal Stromal Cells under Physiological Oxygen Level. Biochemistry Moscow 84, 263–271 (2019). https://doi.org/10.1134/S0006297919030088

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1134/S0006297919030088

Keywords

Search

Navigation