UDC

[611.018.7+611.611]-092.9

DOI

10.17238/issn2542-1298.2019.7.1.16

Authors

Vladimir P. Pashchenko*, Elena V. Tikhonova*, Andrey B. Gudkov*/**
*Northern State Medical University (Arkhangelsk, Russian Federation)
**Northern (Arctic) Federal University named after M.V. Lomonosov (Arkhangelsk, Russian Federation)

Abstract

The method of cell and tissue culture in vitro is widely used for the study of structural cytophysiological properties of body cells. Placing tissue fragments into artificial culture media allows one to assess the viability of tissues as well as the regenerative and adaptive ability of cells. This paper presents the results of quantitative assessments of in vitro growth capacity of kidney tissue fragments from animals (mice) of different ages, using the method of multiple tissue cultures. Renal tissue fragments were cultivated under identical conditions on the surface of perforated Millipore filters. For a single assessment of culture growth, up to 315 tissue microfragments were used, allowing us to perform a more accurate assessment of their quantitative parameters. The growth was estimated by photometry using the Imagej computer program. The research revealed that with ageing the ability of tissues to form growing cell colonies decreases significantly and their growth slows down. The comparison of tissues of animals aged 30 and 140 days showed that 4.7 times less cultures were produced from older animals and their mean area was 1.8 times less than that of cultures from older animals; whereas in cultures from animals aged 10 and 160 days, the difference was 15.7 times and 4.3 times, respectively. The research helped to clarify the pronounced dependence of the growth of epithelial cells of kidney tissue fragments on the age of animals, which is manifested in the decreased number of formed cultures and inhibited growth of cell colonies.

Keywords

tissue growth, tissue culture, regeneration, ageing

References

1. Paul J. Cell and Tissue Culture. Edinburgh, 1960 (Russ. ed.: Pol D. Kul’tura kletok i tkani. Moscow, 1963. 347 p.).
2. Pinaev G.P. Kletochnye kul’tury v fundamental’nykh i prikladnykh issledovaniyakh [Cell Cultures in Basic and Applied Research]. Metody kul’tivirovaniya kletok [Cell Culture Methods]. St. Petersburg, 2008, pp. 7–22.
3. Krylova T.A., Musorina A.S., Zenin V.V., Kol’tsova A.M., Kropacheva I.V., Turilova V.I., Yakovleva T.K., Polyanskaya G.G. Poluchenie i kharakteristika neimmortalizovannykh kletochnykh liniy dermal’nykh fibroblastov cheloveka, vydelennykh iz kozhi vek vzroslykh donorov raznogo vozrasta [Derivation and Characteristic of Non-Immortalized Cell Lines of Human Dermal Fibroblasts, Generated from Skin of the Eyelids of Adult Donors of Different Age]. Tsitologiya, 2016, vol. 58, no. 11, pp. 850–864.
4. Hayflick L. The Limited in vitro Lifetime of Human Diploid Cell Strains. Exp. Cell Res., 1965, vol. 37, no. 3, pp. 614–636.
5. Spasokukotskiy Yu.A., Barchenko L.I. Vliyanie biologicheski aktivnykh veshchestv na protsessy stareniya [The Influence of Biologically Active Substances on the Processes of Ageing]. Biologiya stareniya [The Biology of Ageing]. Leningrad, 1982, pp. 586–600.
6. Nikolenko N.S. Stvolovye kletki vzroslogo organizma v kul’ture [Stem Cells of an Adult Body in vitro]. Metody kul’tivirovaniya kletok [Cell Culture Methods]. St. Petersburg, 2008, pp. 188–201.
7. Pinaev G.P. Problemy i perspektivy razvitiya kletochnykh tekhnologiy [Problems and Prospects of the Development of Cell Technologies]. Kletochnye tekhnologii dlya regenerativnoy meditsiny [Cell Technologies for Regenerative Medicine]. St. Petersburg, 2011, pp. 8–24.
8. Kudryavtseva M.V., Zavadskaya E.E., Skorina A.D., Smirnova S.A., Kudryavtsev B.N. Metod polucheniya izolirovannykh kletok pecheni cheloveka iz materiala prizhiznennykh punktsionnykh biopsiy [The Method of Obtaining Isolated Human Liver Cells from the Material of Intravital Needle Biopsies]. Laboratornoe delo, 1983, no. 9, pp. 21–23.
9. Lopukhin Yu.M., Kogan E.M. Kriterii zhiznesposobnosti organov i tkaney pered transplantatsiey [Criteria of Viability of Organs and Tissues Prior to Transplantation]. Moscow, 1975. 280 p.
10. Khey R. Sokhranenie i otsenka kachestva kletok [Preservation and Quality Assessment of Cells]. Kul’tura zhivotnykh kletok: metody [Culture of Animal Cells: Methods]. Moscow, 1989, pp. 108–165.
11. Pashchenko V.P. Ustroystvo dlya posadki na podlozhku mikrofragmentov biologicheskikh tkaney [Device for Planting Microfragments of Biological Tissues on a Substrate]. Patent USSR no. 1785530, 1991.
12. Schneider C.A., Rasband W.S., Eliceiri K.W. NIH Image to ImageJ: 25 Years of Image Analysis. Nat. Methods, 2012, vol. 9, no. 7, pp. 671–675.
13. Noursadeghi M., Tsang J., Haustein T., Miller R.F., Chain B.M., Katz D.R. Quantitative Imaging Assay for NF-κB Nuclear Translocation in Primary Human Macrophages. J. Immunol. Methods, 2008, vol. 329, no. 1-2, pp. 194–200. DOI: 10.1016/j.jim.2007.10.015
14. Liozner L.D. (ed.). Novoe v uchenii o regeneratsii [New Developments in the Theory of Regeneration]. Moscow, 1977. 358 p.
15. Pashchenko V.P., Balashov V.K., Sivkov A.N., Pashchenko A.V. Ispol’zovanie metoda tkanevykh kul’tur dlya otsenki kachestva pit’evoy vody [Using the Method of Tissue Cultures to Assess Drinking Water Quality]. Ekologiya cheloveka, 2003, no. 3, pp. 27–29.
16. Pashchenko V.P., Gudkov A.B. Vliyanie pit’evoy vody “Akvarel’” na kletki tkani pochek pri kul’tivirovanii in vitro [The Influence of Akvarel Drinking Water on Kidney Cells Cultivated in vitro]. Ekologiya cheloveka, 2006, no. 4 (suppl. 2), p. 396.
17. Zalkind S.Ya. Dinamika tsitologicheskikh izmeneniy kul’tiviruemykh vne organizma kletok v norme i patologii [Dynamics of Cytological Changes in Normal and Pathological Cells Cultivated in vitro]. Voprosy tsitologii, gistologii i embriologii [Issues of Cytology, Histology and Embryology]. Riga, 1960, pp. 15–20.
18. Zalkind S.Ya., Tsypkin L.B. Nekotorye voprosy differentsirovki i adaptatsii v kletochnykh kul’turakh [Some Issues of Differentiation and Adaptation in Cell Cultures]. Tsitologiya, 1971, vol. 13, no. 5, pp. 547–563.
19. Onishchenko N.A. Kletochnye tekhnologii i sovremennaya meditsina [Cell Technologies and Modern Medicine]. Patologicheskaya fiziologiya i eksperimental’naya terapiya, 2004, no. 4, pp. 2–11.