Abstract
The studies were conducted on 2-3-months-old males of C57BL/6 mice weighing 20–24 g. Obtaining and cultivating of adipose-derived mesenchimal stem cells (AD MSCs) were carried out in a sterile laminar box with compliance of conditions of asepsis and antiseptics. AD MSCs of the 2, 4, 7 and 12 passages were analyzed. Morphometric analysis was performed using a light microscopy. Morphometric parameters such as cell and nucleus area or nuclear-cytoplasmic ratio (NCR) were calculated using the Axiovision light microscope (Carl Zeiss, Germany) and ImageJ 1.45 software. Trypan blue dye used for investigation of the viability of MSC. The morphological characteristics of mesenchymal stem cells from adipose tissue during the process of cultivation changes: at the first passages of cultivation, the cells are spindle-shaped with two, at least three, long long cytoplasmic processes, located bipolar. Near the nucleus the Golgi complex is clearly visible – a sign of active cells. At later passages cells have a small cytoplasmic processes and the bipolar arrangement of processes changes by stellar arrangement. Golgi complex is also clearly visualized. The indicator of the nuclear-cytoplasmic ratio in MSC from adipose tissue is significantly reduced at 7 passage to 0.2189 ± 0.0122 (P < 0.01), and at 12 passage to 0.1111 ± 0.0086 (P < 0.001) compared to the 2 passage. The coefficient of proliferation of MSC from adipose tissue is significantly reduced at 12th passage. The viability of mesenchymal stem cells from adipose tissue with an increasing of a number of passages significantly reduces and at the 12th passage of cultivation reaches 84,67 ± 1,36* (P < 0.05). The content of apoptotic cells that exhibited sensitivity to serum-free significantly increased at 7 and 12 passages and was respectively 21.33 ± 1.36 (P < 0.05) and 23.67 ± 0.97% (P < 0.05).
Highlights
It is known that mesenchymal stem cells in the bone marrow make up 0.001% to 0.01% of the total fraction of mononuclear cells, and bone marrow aspiration is an invasive procedure and has a significant effect on the donor after the surgical period (Dmitrieva et al, 2012)
The fatty tissue is an excellent alternative source of mesenchymal stem cells, since it contains approximately 500 times more MSC compare to bone marrow
Near the nucleus in zone of enlightenment the Golgi complex is clearly defined, which is well developed in active forms of cells
Summary
It is known that mesenchymal stem cells in the bone marrow make up 0.001% to 0.01% of the total fraction of mononuclear cells, and bone marrow aspiration is an invasive procedure and has a significant effect on the donor after the surgical period (Dmitrieva et al, 2012). Other sources of stem cells, in particular umbilical cord blood, placenta, are used in modern medicine and veterinary medicine (Ning et al, 2012). The fatty tissue is an excellent alternative source of mesenchymal stem cells, since it contains approximately 500 times more MSC compare to bone marrow. Some data are already known about biological properties of adipose derived MSC (aMSC). It is known about high differential potential of MSC from fatty tissue of animals of different species, their immunomodulatory property. Some authors emphasize that they exhibit stronger immunomodulatory effects, due to the fact that
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