ISSN 1990-7478, Biochemistry (Moscow) Supplement Series A: Membrane and Cell Biology, 2007, Vol. 1, No. 4, pp. 284–293. © Pleiades Publishing, Ltd., 2007. Original Russian Text © M.V. Moldaver, E.B. Dashinimaev, K.S. Vishnyakova, P.M. Chumakov, Y.E. Yegorov, 2007, published in Biologicheskie Membrany, 2007, Vol. 24, No. 5, pp. 402–412. 284 Numerous studies are devoted to applications of cell technologies in medicine nowadays. Transplantation of autologous fibroblasts and mesenchymal stem cells is now performed for treatment of skin lesions [1]. Stem cells are likely to find application in therapy of such dis- eases as myocardial infarction, osteochondrosis, neuro- degenerative syndromes, ischemia, and other condi- tions associated with tissue disorders caused by cell degradation [2]. An average human life span has increased consider- ably over the last decades in developed countries. Human aging is related to the absence of the sufficient amount of cells capable to fully compensate for a natu- ral loss of cells; this misbalance results in degenerative syndromes. In this period the number of stem and pro- genitor cells decreases, and the lack of sufficient amounts of cells make it impossible to use them for transplantation therapy. Harvesting of cells from primary cultures and cell culturing in vitro are also coupled with various difficul- ties that go far beyond selection of nutrient media and substrates. Atmospheric air contains much more oxy- gen (21%) than body tissues. Oxygen level in arterial blood is about 15%, in venous blood, 5%, and in carti- laginous tissue, 1%. Increased oxygen level in vitro can differently affect cell behavior. Oxygen effects are manifested in different ways: (i) changes in the protein functional activities due to alterations in protein thiol groups; (ii) alterations of gene expression due to the oxygen-sensitive transcrip- tion factor HIF; (iii) toxic (or signaling?) effects of free radicals; (iv) changes in the oxidative phosphorylation, e.g., in glycolysis ratio, etc. [3, 4]. The most evident influence of atmospheric oxygen on cell cultures is an inhibition of cell proliferation ability. This inhibition is considered to be a result of the telomere damage pro- duced by reactive oxygen species [5], the main source of which is mitochondria. The higher the activity of the electron transport chain, the more intense is the release (leakage) of reactive oxygen species [6]. Under certain conditions, this leakage should correlate with the con- centration of dissolved oxygen. In 1998 it was found that introduction of the telom- erase reverse transcriptase gene (hTERT) into cells results in their immortalization, since the cells acquire an ability to perform an infinite number of population doublings [7, 8] without any side effects of telomerase expression [9, 10]. Cultured cells retained all mecha- nisms of regulation of cell proliferation over hundreds of population doublings (PD) without any signs of malignant transformation. The only reason that restrained telomerized cell application in medicine was Influence of Oxygen on Three Different Types of Telomerized Cells Derived from a Single Donor M. V. Moldaver a , E. B. Dashinimaev b , K. S. Vishnyakova a , P. M. Chumakov a , and Y. E. Yegorov a a Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, ul. Vavilova 32, Moscow, 119991 Russia; e-mail: yegorov@eimb.ru b Moscow Institute of Physics and Technologies, Institutskii per. 9, Dolgoprudnyi, Moscow oblast, 141700 Russia Received April 11, 2007 Abstract—Three primary cell cultures were derived from a single donor: skin fibroblasts (SF), dermal papilla cells (DP), and mesenchimal stromal cells from lipoaspirate (LA). After a high efficiency introduction of the gene of human telomerase catalytic component (hTERT) in lentiviral construct, we obtained three different strains of immortalized cells. All cells were cultured in a low oxygen (3%) environment. When telomerized cells were grown in atmospheric conditions (21% oxygen), growth retardation was observed after a period of 18–40 days. SF-hTERT and DP-hTERT overcame this retardation. In 30–45 days the rate of their growth became as high as it was in 3% oxygen. LA-hTERT cells were incapable to restore the previous growth rate and after several passages ceased proliferation. We found that telomerized cells were heterogeneous in their ability to form colonies at low density (3 cells per 1 cm 2 ): they made 0–9 population doublings in 7 days. At the same time, mean growth rate of SF-hTERT did not change, while the growth of DP-hTERT and LA-hTERT cells was considerably accelerated in comparison with their growth in mass culture. Transfer of telomerized cells to 21% oxygen reduced their ability to form colonies to a different extent. The total number of cells grown in 21% oxy- gen was 3 times less than the number grown in 3% oxygen for SF-hTERT and DP-hTERT, and 6 times less for LA-hTERT. Original cells (with the exception of LA) were more sensitive to oxygen level than the correspond- ing telomerized cells. The total number of cells of all three strains (SF, DP, LA) grown in 21% oxygen was 6– 7 times less. We discuss the reasons of incapability of LA-hTERT cells to adapt to atmospheric oxygen. DOI: 10.1134/S1990747807040034