Medical Radiology and Radiation Safety. 2023. Vol. 68. № 1

DOI: 10.33266/1024-6177-2023-68-1-5-14

V.A. Nikitina, T.A. Astrelina, V.Yu. Nugis, I.V. Kobzeva, E.E. Lomonosova,
Yu.B. Suchkova, T.F. Malivanova, V.A. Brunchukov, D.Yu. Usupzhanova,
V.A. Brumberg, A.A. Rastorgueva, E.I. Dobrovolskaya, T.V. Karaseva,
M.G. Kozlova, M.V. Pustovalova, A.K. Chigasova, N.Yu. Vorobyeva,
A.N. Osipov, A.S. Samoilov

Cytogenetic Analysis of the Cell Line of Multipotent Human Mesenchymal Stromal Cells during Long-Term Cultivation after Exposure to X-Ray Radiation at Low and Medium Doses

A.I. Burnazyan Federal Medical Biophysical Center, Moscow, Russia

Contact person: V.A. Nikitina, e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.

 

ABSTRACT

Purpose: To evaluate the frequency and spectrum of chromosome aberrations under X-Ray exposure at doses of 80, 250, and 1000 mGy in a human multipotent mesenchymal stromal cell (MMSC) cell line during long-term cultivation.

Material and methods: MMSCs were isolated from human gingival mucosa by an enzymatic method and cultured in a serum-free medium. The presence of surface antigens was determined using the method of flow cytometry. The ability of the cell line to differentiate in the osteogenic, adipogenic, and chondrogenic directions was studied using induction media. Authentication was performed by genotyping of polymorphic STR loci, cytogenetic analysis was performed by multicolor fluorescent in situ hybridization (mFISH). Irradiation was carried out on an X-ray biological unit RUB RUST-M1 (Russia) at a dose rate of 40 mGy/min, a voltage of 100 kV, and a current of 0.8 mA.

Results: At the first passage after irradiation, a statistically significant increase in the frequency of non-clonal CA compared with the control was recorded at a dose of 80, but not 250 and 1000 mGy. At the late stages of cultivation, the average frequency of breaks per chromosome in the group of non-irradiated cells did not differ from the values obtained after irradiation at doses of 80, 250, and 1000 mGy (p > 0.05). However, in MMSCs irradiated at a dose of 80 mGy, damage occurred more often in pairs of chromosomes 6 and 10, and at a dose of 1000 mGy, in a pair of chromosomes 9. A single irradiation of MMSCs in vitro did not affect the growth and progression of MMSCs characteristic of the studied primary cell line, of clonal cells with chromosome translocations and monosomy X, but led to an increase in the representation of a clone with tetrasomy 8. The total number of random clones with chromosome translocations that arose de novo increased after irradiation at a dose of 1000 mGy.

Conclusion: Minor fluctuations in the proportion of cells with non-clonal CA, depending on the dose received in the early stages after irradiation (passage 1–4), disappeared at the later stages of cultivation (passage 8–14). There were no differences in mean frequencies between irradiated and non-irradiated MMSCs, but after irradiation, damage to some chromosomes could occur more frequently than others. A single X-ray irradiation of MMSCs can promote the growth and progression of primary pathological cytogenetic clones, regardless of the dose received, as well as an increase in the total number of de novo cell clones with chromosomal translocations that have arisen. A single X-ray irradiation of MMSCs can promote the growth and progression of primary pathological cytogenetic clones, regardless of the dose received, as well as an increase in the total number of de novo cell clones with chromosomal translocations that have arisen.

Keywords: mesenchymal multipotent stromal cells, chromosome aberrations, mFISH, X -ray irradiation, low doses

For citation: Nikitina VA, Astrelina TA, Nugis VYu, Kobzeva IV, Lomonosova EE, Suchkova YuB, Malivanova TF, Brunchukov VA, Usupzhanova DYu, Brumberg VA, Rastorgueva AA, Dobrovolskaya EI, Karaseva TV, Kozlova MG, Pustovalova MV, Chigasova AK, Vorobyeva NYu, Osipov AN, Samoilov AS. Cytogenetic Analysis of the Cell Line of Multipotent Human Mesenchymal Stromal Cells during Long-Term Cultivation after Exposure to X-Ray Radiation at Low and Medium Doses. Medical Radiology and Radiation Safety. 2023;68(1):5–14. (In Russian). DOI: 10.33266/1024-6177-2023-68-1-5-14

 

References

1. Niwa O., Barcellos-Hoff M.H., Globus R.K., Harrison J.D., Hendry J.H., Jacob P., et al. ICRP Publication 131: Stem Cell Biology with Respect to Carcinogenesis Aspects of Radiological Protection. Ann. ICRP. 2015;44;3-4:7-357. DOI: 10.1177/0146645315595585.

2. Hendry J.H., Niwa O., Barcellos-Hoff M.H., Globus R.K., Harrison J.D., Martin M.T., et al. ICRP Publication 131: Stem Cell Biology with Respect to Carcinogenesis Aspects of Radiological Protection. Ann. ICRP. 2016;45;1:239-252. DOI: 10.1177/0146645315621849.

3. Morikawa S., Mabuchi Y., Kubota Y., Nagai Y., Niibe K., Hiratsu E., et al. Prospective Identification, Isolation, and Systemic Transplantation of Multipotent Mesenchymal Stem Cells in Murine Bone Marrow. J. Exp. Med. 2009;206;11:2483-2496. DOI: 10.1084/jem.20091046.

4. Cairns J. Mutation Selection and the Natural History of Cancer. Nature. 1975;255:197–200. DOI: 10.1038/255197a0.

5. Ilin L.A., Rozhdestvenskiy L.M., Koterov A.N., Borisov N.M. Aktualnaya Radiobiologiya = Actual Radiobiology. A Course of Lectures. Moscow Publ., 2015. 240 p. ISBN 978-5-383-00932-1 (In Russ.).

6. Gorbunova V.N., Baranov V.S. Vvedeniye v Molekulyarnuyu Diagnostiku i Genoterapiyu Nasledstvennykh Zabolevaniy = Introduction to Molecular Diagnostics and Gene Therapy of Hereditary Diseases. A Textbook for Students of Medical Universities. St. Petersburg Publ., 1997. 287 p. ISBN 5-87685-076-4 (In Russ.).

7. Gothe H.J., Minneker V., Roukos V. Dynamics of Double-Strand Breaks: Implications for the Formation of Chromosome Translocations. Adv. Exp. Med. Biol. 2018;1044:27-38. DOI:10.1007/978-981-13-0593-1_3.

8. Terskikh V.V., Vasilyev A.V., Vorotelyak Ye.A. Polarization and Asymmetric Division of Stem Cells. Tsitologiya. 2007;49;11:933-938 (In Russ.).

9. Bochkov N.P., Nikitina V.A. Cytogenetics of Human Stem Cells. Molekulyarnaya Meditsina = Molecular Medicine. 2008;3:40-47 (In Russ.).

10. Chen M.F., Lin C.T., Chen W.C., Yang C.T., Chen C.C., Liao S.K., et al. The Sensitivity of Human Mesenchymal Stem Cells to Ionizing Radiation. Int. J. Radiat. Oncol. Biol. Phys. 2006;66;1:244-253. doi:10.1016/j.ijrobp.2006.03.062.

11. Fekete N., Erle A., Amann E.M., Fürst D., Rojewski M., Langonné A., et al. Effect of High-Dose Irradiation on Human Bone-Marrow-Derived Mesenchymal Stromal Cells. Tissue Engineering Part C Methods. 2015;21;2:112-122. DOI: 10.1089/ten.TEC.2013.0766. 

12. Nicolay N.H., Lopez Perez R., Saffrich R., Huber P.E. Radio-Resistant Mesenchymal Stem Cells: Mechanisms of Resistance and Potential Implications for the Clinic. Oncotarget. 2015;6;23:19366-19380. DOI: 10.18632/oncotarget.4358. 

13. Sugrue T., Lowndes N.F., Ceredig R. Mesenchymal Stromal Cells: Radio-Resistant Members of the Bone Marrow. Immunol Cell. Biol. 2013;91;1:5-11. DOI: 10.1038/icb.2012.61. 

14. Rieger K., Marinets O., Fietz T., Körper S., Sommer D., Mücke C., et al. Mesenchymal Stem Cells Remain of Host Origin Even a Long Time after Allogeneic Peripheral Blood Stem Cell or Bone Marrow Transplantation. Exp. Hematol. 2005;33;5:605-611. doi: 10.1016/j.exphem.2005.02.004.

15. Lomonosova Ye.Ye., Nugis V.Yu., Snigireva G.P., Kozlova M.G., Nikitina V.A., Galstyan I.A. Cytogenetic Analysis of Peripheral Blood Lymphocyte Cultures of the Patient in the Long Term after Emergency Irradiation Using the Tricolor FISH Method. Radiatsionnaya Biologiya. Radioekologiya = Radiation Biology. Radioecology. 2022;62;1:5-17. DOI: 10.31857/S0869803122010064 (In Russ.).

16. Dominici M., Le Blanc K., Mueller I., Slaper-Cortenbach I., Marini F., Krause D., et al. Minimal Criteria for Defining Multipotent Mesenchymal Stromal Cells. The International Society for Cellular Therapy Position Statement. Cytotherapy. 2006;8;4:315-317. doi: 10.1080/14653240600855905.

17. Shaffer L.G., McGowan-Jordan J., Schmid M. ISCN 2013: an International System for Human Cytogenetic Nomenclature - 2013. Basel, Karger, 2013.

18. Nikitina V.A., Astrelina T.A., Kobzeva I.V., Nugis V.Yu., Lomonosova Ye.Ye., Semina V.V., et al. The Cytogenetic Characteristic of Mesenchymal Multipotent Stromal Cell Diploid Lines. Tsitologiya. 2021;63;3:207-220. DOI 10.31857/S0041377121030081 (In Russ.).

19. Bochkov N.P., Voronina Ye.S., Katosova L.D., Nikitina V.A. Cytogenetic Study of Multipotent Mesenchymal Stromal Cells of Humans in the Process of Cultivation. Meditsinskaya Genetika = Medical Genetics. 2009;12;90:3-6 (In Russ.).

20. Koltsova A.M., Zenin V.V., Petrosyan M.A., Turilova V.I., Yakovleva T.K., Polyanskaya G.G. Isolation and Characterization of Mesenchymal Stem Cells Derived from Different Regions of the Placenta of the Same Donor. Tsitologiya. 2020;62;9:623-637. DOI:10.31857/S0041377120090035 (In Russ.).

21. Polyanskaya G.G. Comparative Analysis of the Lines of human Mesenchymal Stem Cells Derived in the Collection of Cell Cultures of Vertebrates. (Review). Kletochnyye Kultury. 2018;34:3-18 (In Russ.).

22. Barkholt L., Flory E., Jekerle V., Lucas-Samuel S., Ahnert P., Bisset L., et al. Risk of Tumorigenicity in Mesenchymal Stromal Cell-Based Therapies - Bridging Scientific Observations and Regulatory Viewpoints. Cytotherapy. 2013;15;7:753-759. DOI:10.1016/j.jcyt.2013.03.005. 

23. Pustovalova M., Grekhova A., Astrelina Т., Nikitina V., Dobrovolskaya E., Suchkova Y., et al. Accumulation of Spontaneous γH2AX Foci in Long-Term Cultured Mesenchymal Stromal Cells. Aging. 2016;8;12:3498-3506. DOI: 10.18632/aging.101142.

24. Pustovalova M., Astrelina Т.A., Grekhova A., Vorobyeva N., Tsvetkova A., Blokhina T., et al. Residual γH2AX Foci Induced by low Dose X-Ray Radiation in Bone Marrow Mesenchymal Stem Cells Do Not Cause Accelerated Senescence in the Progeny of Irradiated Cells. Aging. 2017;9;11:2397-2410. DOI: 10.18632/aging.101327.

25. Nikitina V., Nugis V., Astrelina T., Zheglo D., Kobzeva I., Kozlova M., et al. Pattern of Chromosomal Aberrations Persisting Over 30 Years in a Chernobyl Nuclear Power Plant Accident Survivor: Study Using mFISH. J. Radiat. Res. 2022;63;2:202-212. DOI:10.1093/jrr/rrab131.

26. Velichko A.K., Razin S.V., Kantidze O.L. Cellular Response to DNA Damage Occurring in Ribosomal Genes. Molekulyarnaya Biologiya = Molecular Biology. 2021;55;2:210-222. DOI: 10.31857/S0026898421020142 (In Russ.).

27. Hemsing A.L., Hovland R., Tsykunova G., Reikvam H. Trisomy 8 in Acute Myeloid Leukemia. Expert Rev. Hematol. 2019;12;11:947-958. DOI: 10.1080/17474086.2019.1657400.

28. Dugan L.C., Bedford J.S. Are Chromosomal Instabilities Induced by Exposure of Cultured Normal Human Cells to Low- or High-LET Radiation? Radiat. Res. 2003;159;3:301-311. DOI:10.1667/0033-7587(2003)159[0301:aciibe]2.0.co;2.

29. Serakinci N., Guldberg P., Burns J.S., Abdallah B., Schrødder H., Jensen T., et al. Adult Human Mesenchymal Stem Cell as a Target for Neoplastic Transformation. Oncogene. 2004;23;29:5095-5098. DOI:10.1038/sj.onc.1207651.

30. Nikitina V., Astrelina T., Nugis V., Ostashkin A., Karaseva T., Dobrovolskaya E., et al. Clonal Chromosomal and Genomic Instability During Human Multipotent Mesenchymal Stromal Cells Long-Term Culture. PLoS One. 2018;13;2:e0192445. DOI:10.1371/journal.pone.0192445.

 

 PDF (RUS) Full-text article (in Russian)

Conflict of interest. The authors declare no conflict of interest.

Financing. The study had no sponsorship.

Contribution. Article was prepared with equal participation of the authors.

Article received: 20.09.2022. Accepted for publication: 25.11.2022.