Ali S. G., Kovalenko I. F., Bozhok G. A.

BIOLOGY

Scentific article

Cell culture of dorsal root ganglia (DRG) is a modern scientific platform for studying the mechanisms of neurogenesis, neurotransmission, neuroregeneration, as well as a valuable source of neural stem cells for regenerative medicine. Goal. To study the expression of b-III-tubulin, glutamine synthetase, S100 protein and chromogranin A in the dorsal root ganglia cell culture of neonatal piglets by immunocytochemical method. Methods. Cell suspension was obtained enzymatically from DRG of new-born piglets. Cells were seeded at a concentration of 5×105 cell/ml on plastic Petri dishes with a surface covered by poly-D-lysine, and cultured in a nutrient medium α-MEM with the addition of 10% fetal calf serum (FCS). After reaching the confluent monolayer, the cells of the primary culture were detached from the surface with a trypsin-EDTA mixture, washed by centrifugation, diluted to the above concentration and subcultured in the same conditions. The cell monolayer formed on day 7 of second passage was subjected to immunocytochemical labeling with antibody against neuronal marker b-III-tubulin, neuroglia markers glutamine synthetase (GS) and S100 protein, as well as neuroendocrine cell marker chromoranin A (ChrA). Results. The immunocytochemical labeling showed a phenotypic heterogeneity of the culture. Cells expressing b-III-tubulin differed morphologically. Some of them were small with long branching processes that form a network, so they were identified as neurons. Others were large elongated polygonal cells with small processes. Small spindle-shaped and large polygonal cells expressed GS, which characterized them as satellite glial cells (SGC). They represented the main cell mass of the obtained culture. S100-positive cells were not observed in the culture. A certain number of ChrA-positive cells were present in the culture. Morphologically, they were cells of various sizes and shapes, among which cells with processes prevailed. Conclusions. The results of the study allow us to conclude that neonatal piglets’ DRG derived cell culture obtained in conditions of poly-D-lysine growth surface and 10% FCS supplemented nutrient medium is heterogeneous in phenotypic composition. The main cellular subpopulations in it are neurons and SGC. This allows us to recommend the obtained culture as a valuable model containing a combination of neurons and glial cells, physiologically close to observed in the body.

dorsal root ganglia, cell culture, neurons, satellite glial cells, b-III-tubulin, glutamine synthetase, chromogranin A.

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«Bulletin of problems biology and medicine» Issue 3 (152), 2019 year, 46-50 pages, index UDK 611.018.82.085.2“461”

10.29254/2077-4214-2019-3-152-46-50