Comparative analysis of genetic stability of rat adipose and bone marrow cell cultures on earley passages

  • V.V. Kovpak National University of life and environmental sciences of Ukraine, Polkovnyka Potekhyna Str., 16, Kyiv, 03041
  • O.S. Kovpak National University of life and environmental sciences of Ukraine, Polkovnyka Potekhyna Str., 16, Kyiv, 03041
Keywords: cytogenetic analysis, micronucleus test, mitotic index, apoptotic index, adipose tissue cell culture, bone marrow cell culture


From literary sources, we received a number of controversial data regarding the risks of neoplastic transformation of stem cells in vitro. Inasmuch as today bone marrow has been sufficiently studied as a source of adult donor stem cells, and adipose tissue is an alternative source for obtaining cell material (from which it may be obtained using less invasive methods and in much greater quantity), determination of genetic stability of these particular cell cultures currently is the most relevant issue. Comparison of genetic stability of rat bone marrow and adipose cell cultures from the first to the sixth passages. Materials and methods: 3 non-linear 4 months old male rats and 9 non-linear 12 days old infant rats were used in the experiment. Adipose cell culture was obtained from subcutaneous fat using the standardized method in our own modification. Bone marrow cell culture was obtained from the rat femurs, tibiae, and humeri using the standardized method. Obtained cell mass was cultivated in a standard culture medium in a СО2 incubator at 37 ºС and with 5% СО2 concentration. Cytogenetic analysis was carried out on 30 metaphase plates from each passage. Cells from the first to the sixth passage we used in the research. A modification of the standard cytogenetic method was used to obtain chromosome preparations. In the preparations prepared in an aforementioned way, numerical chromosome abnormalities, like aneuploidy, polyploidy, were determined, as well as the number of binucleate cells, micronucleus cells, mitotic and apoptotic indexes. Here, we provide the comparison of genetic stability indicators for rat bone marrow and adipose cell cultures throughout the in vitro cultivation thereof. Changes in the genetic apparatus were identified in both bone marrow and adipose tissue cell cultures. Karyotype analysis of rat bone marrow and adipose cell cultures showed that, given the conditions that we used for the cultivation, the number of aneuploidies and polyploidies varies from one passage to another; however it does not go beyond the limits of spontaneous mutagenesis typical for mammals. Based on the results of cytogenetic evaluation of the culture, it was determined that the number of cells with micronuclei and the binucleate cell remains within the normal range from the first to the sixth passages. The number of cells with aneuploidy and micronuclei in adipose cell culture on all passages was lower than in bone marrow cell culture; this indicates its higher genetic stability in terms of these indicators. The highest mitotic index was determined on the first passage with its following decrease throughout the cultivation in inverse proportion to apoptotic index. The obtained data on the genetic stability of both adipose and bone marrow cell cultures are within normal range typical for mammals in terms of all studied indicators. This allows for the following studies relating to the transplantation with minimal risk of neoplastic transformation of the said cultures.


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Bersenev, A.B. (2005). Izucheniye spontannoy onkogeneticheskoy transformatsii mezenkhimal'nykh stvolovykh kletok cheloveka v kul'ture [Study of spontaneous oncogenetic transformation of human mesenchymal stem cells in culture] Kletochnaya transplantologiya i tkanevaya inzheneriya.– Cellular transplantation and tissue engineering. 1, 14–16 (in Russian).

Kolmakowa, T.C., Belik, C.N., Morgul', E.W., Cewrjukow, A.W. (2013). Icpol'sowanie mikrojadernogo tecta dlja ozenki jevvektiwnocti letschenija allergii u detej: metod. rekomendazii. [Using the micronucleus test to assess the effectiveness of the treatment of allergies in children: guidelines]. Roctow na Donu. Isd-wo RoctGMU (in Russian).

Kovaljowa, O., Koboseva, N., Burdo, E., Glasko, T. (2008). Mikrojadernyj tect kak metod opredelenija cesonnoj ismentschiwocti zitogenetitscheckich pokasatelej u mlekopetajushich [Micronucleus test as a method of determining the seasonal variability of cytogenetic indices in of mammals]. Rarіtetna terіovauna ta її ochorona – Rare fauna and its protection. 9, 266–269 (in Ukrainian).

Kovaljova, O.A. (2008). Zitogenetitscheckie anomalii w comatitscheckich kletkach mlekopitajushich. [Cytogenetic abnormalities in somatic cells at mammalian]. Zitologija i genetika – Cytology and Genetics. 1, 58–72 (in Ukrainian).

Mazurkevych, A.Y., Malyuk, M.O., Starodub, L.F., Danilov, V.B. (2015). Kariotypovyy analiz mezenkhimalʹnykh stovburovykh klityn kistkovoho mozku kroliv za riznykh metodiv dysotsiatsiyi klitynnoho monosharu na rannikh pasazhakh kulʹtyvuvannya in vitro [Kariotyp analysis of mesenchimal bone marrow stem cells of rabbits by different methods dissociation of cell monolayers at early cultivation passage in vitro]. Naukovyy visnyk NUBiP Ukrayiny. Seriya: Veterynarna medytsyna, yakistʹ i bezpeka produktsiyi tvarynnytstva – Scientific Bulletin NUBIP of Ukraine. Series: Veterinary Medicine, the quality and safety of animal products, 227 (in Ukrainian).

Mazurkevych, A.Y., Kovpak, V.V., Kovpak, O.S. (2016). Fenotypovi ta morfolohichni zminy kulʹtury klityn kistkovoho mozku shchuriv v protsesi yikh kulʹtyvuvannya [Phenotypic and morphological changes of bone marrow cells culture of rats during cultivation]. Naukovyy visnyk LNUVMBT imeni S.Z.Gzhytsʹkoho – Scientific Messenger LNUVMBT named after S.Z. Gzhytskyj. 18, 2(66), 126–131 (in Ukrainian).

Masurkewitsch, A.J., Kowpak, W.W., Danіlow, W.B. (2014). Klіtinnі technologії u weterinarnіj medizinі. Nawtschal'nij pocіbnik. [Cellular technologies in veterinary medicine. Study Guide]. Kyev: KOMPRINT (in Ukrainian).

Muschkambarow, N.N., Kusnezow, C.L. (2013). Molekuljarnaja biologija.Utschebnoe pocobie dlja ctudentow medizinckich wusow [Molecular biology. Textbook for medical students]. Moscow: Medizinckoe invormazionnoe agentctwo (in Russian).

Jernct, L.K., Zhigatschew, A.I. (2006). Monitoring genetitscheckich bolesnej zhiwotnych w cicteme krupnomacschtabnoj celekzii [Monitoring animal genetic diseases in the system of largescale selection]. Moscow: Roccel'chosakademija (in Russian).

Bunnell, B.A., Flaat, M., Gagliardi, Ch. et al. (2008). Adipose-derived stem cells: isolation, expansion and differentiation. Methods. 45, 2, 115–120.

Carswell, K.A., Mi-Jeong, Lee, Fried, S.K. (2012). Culture of Isolated Human Adipocytes and Isolated Adipose Tissue. Methods Mol Biol. 806, 203–214.

Foudah, D., Redaelli, S., Donzelli, E. et al. (2009). Monitoring the genomic stability of in vitro cultured rat bone-marrow-derived mesenchymal stem. Chromosome Res.17(8), 1025–1039.

Ian Freshney, R. (2005). Culture of animal cells: a manual of basic technique [5th ed.]. USA: John Wiley & Sons.

Izadpanah, R., Kaushal, D., Kriedt, C., Tsien, F. et al. (2008). Long-term In vitro Expansion Alters the Biology of Adult Mesenchymal Stem. Cancer Res. 68(11), 4229–4238.

Mareschi, K., Ferrero, I., Rustichelli, D., Aschero, S. et al. (2006). Expansion of mesenchymal stem cells isolated from pediatric and adult donor bone marrow. J. Cell. Biochem. 97(4), 744–754.

Matsyara, T. (1980) Multinucleation and polypoloidization of aging human cells in culture. Adv Exp Med Biol. 129, 31–38.

Miura, M., Miura, Y., Padilla, N.M. et al (2005). Accumulated chromosomal instability in murine bone marrow mesenhymal stem cells leads to malignant transformation. Stem. Cells. 24, 1095–1103.

Penaloza, C., Orlanski, S., Entezari-Zaher, T. et al. (2008). Cell death in mammalian development. Curr. Pharm. Des. 14(2), 184–196.

Rubio, D., Garcia Castro, J., Martin, M.C. et al. (2005) Spontaneous human stem cell trans formation. Cancer Res. 65, 3035–3039.

Xikum, X., Liming, S. (1990). Observations on micronuclei germ cells. Zool. Res. 11, 4, 343.

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Kovpak, V., & Kovpak, O. (2017). Comparative analysis of genetic stability of rat adipose and bone marrow cell cultures on earley passages. Scientific Messenger of LNU of Veterinary Medicine and Biotechnologies. Series: Veterinary Sciences, 19(73), 95-100.