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


About the author:

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



Type of article:

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 5x105 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.


  1. Malin SA, Davis BM, Molliver DC. Production of dissociated sensory neuron cultures and considerations for their use in studying neuronal function and plasticity. Nat Protoc. 2007;2(1):152-60. DOI: 10.1038/ nprot.2006.461
  2.  Wong AW, KP Yeung J, Payne SC, Keast JR, Osborne PB. Neurite outgrowth in normal and injured primary sensory neurons reveals different regulation by nerve growth factor (NGF) and artemin. Mol Cell Neurosci. 2015;65:125-34. DOI: 10.1016/j.mcn.2015.03.004 PMID: 25752731
  3. Muratori L, Ronchi G, Raimondo S, Geuna S, Giacobini-Robecchi MG, Fornaro M. Generation of new neurons in dorsal root ganglia in adult rats after peripheral nerve crush injury. Neural Plast. 2015. 860546. DOI: 10.1155/2015/860546
  4. Ciaroni S, Cecchini T, Cuppini R, Ferri P, Ambrogini P, Bruno C, et al. Are there proliferating neuronal precursors in adult rat dorsal root ganglia? Neurosci Lett. 2000;281(1):69-71.
  5. Li HY, Say EH, Zhou XF. Isolation and characterization of neural crest progenitors from adult dorsal root ganglia. Stem Cells. 2007;25(8): 2053-65.
  6. Singh RP, Cheng YH, Nelson P, Zhou FC. Retentive multipotency of adult dorsal root ganglia stem cells. Cell Transplant. 2009;18(1):55-68.
  7. Meller K. The Reaggregation of Neurons and Their Satellite Cells in Cultures of Trypsin-Dissociated Spinal Ganglia. Cell Tiss. Res. 1974;152: 175-83.
  8. Mudge AW. Effect of non-neuronal cells on peptide content of cultured sensory neurones. J Exp Biol. 1981;95:195-203.
  9. de Luca AC, Faroni A, Reid AJ. Dorsal root ganglia neurons and differentiated adipose-derived stem cells: An in vitro co-culture model to study peripheral nerve regeneration. J Vis Exp. 2015;96: e52543. [Cited 28.07.2019]. Available from: PMC4354675/
  10. Huang T, Cherkas P, Rosenthal D. Dye coupling among satellite glial cells in mammalian dorsal root ganglia. Brain Res Brain Res Rev. 2005;1036(1-2):42-9.
  11. Backström E, Chambers BJ, Kristensson K, Ljunggren HG. Direct NK cell-mediated lysis of syngenic dorsal root ganglia neurons in vitro. J Immunol. 2000;165(9):4895-900.
  12. Svenningsen A, Colman DR, Pedraza L. Satellite cells of dorsal root ganglia are multipotential glial precursors. Fex Neuron Glia Biol. 2004;1(1):85-93.
  13. Gerhauser I, Hahn K, Baumgärtner W, Wewetzer K. Culturing adult canine sensory neurons to optimise neural repair. Vet Rec. 2012;170(4):102.
  14. Morgan BR, Coates JR, Johnson GC. Characterization of thoracic motor and sensory neurons and spinal nerve roots in canine degenerative myelopathy, a potential disease model of amyotrophic lateral sclerosis. J Neurosci Res. 2014;92(4):531-41. DOI: 10.1002/jnr.23332
  15.  Fadda A, Bärtschi M, Hemphill A, Widmer HR, Zurbriggen A, Perona P, et al. Primary postnatal dorsal root ganglion culture from conventionally slaughtered calves. PLoS One. 2016;11(12):e0168228. DOI: 10.1371/journal.pone.0168228
  16. Tongtako W, Lehmbecker A, Wang Y, Hahn K, Baumgärtner W, Gerhauser I. Canine dorsal root ganglia satellite glial cells represent an exceptional cell population with astrocytic and oligodendrocytic properties. Sci Rep. 2017;7(1):13915.
  17. Bassols A, Costa C, Eckersall PD, Osada J, Sabrià J, Tibau J. The pig as an animal model for human pathologies: a proteomics perspective. Proteomics Clin. Appl. 2014;8(9):715-31.
  18. Ali SG, Sidorenko OS, Bozhok GA. Vliyaniye sostava pitatel’noy sredy na morfologicheskiye kharakteristiki kul’tury kletok spinal’nykh gangliyev neonatal’nykh porosyat. Visnyk Kharkivsʹkoho natsionalʹnoho universytetu imeni V.N. Karazina. Seriya «Biolohiya». 2018;30:49-59. DOI: 10.26565/2075-5457-2018-30-6 [in Russiаn].
  19. Delree P, Leprince P, Schoenen J, Moonen G. Purification and culture of adult rat dorsal root ganglianeurons. J Neurosci Res. 1989;23(2): 198-206.
  20. Liu R, Lin G, Xu H. An efficient method for dorsal root ganglia neurons purification with a one-time anti-mitotic reagent treatment. PLoS One. 2013;8(4):e60558. DOI: 10.1371/journal.pone.0060558
  21.  Chen ZL, Strickland S. Laminin gamma1 is critical for Schwann cell differentiation, axon myelination, and regeneration in the peripheral nerve. J Cell Biol. 2003;163(4):889-99.
  22. Haastert K, Mauritz C, Chaturvedi S, Grothe C. Human and rat adult Schwann cell cultures: fast and efficient enrichment and highly effective non-viral transfection protocol. Nat Protoc. 2007;2(1):99-104.
  23. Marin V, Kaplanski G, Grès S, Farnarier C, Bongrand P. Endothelial cell culture: protocol to obtain and cultivate human umbilical endothelial cells. Journal of Immunological Methods. 2001;254(1-2):183-90. DOI: 10.1016/s0022-1759(01)00408-2
  24. Belzer V, Shraer N, Hanani M. Phenotypic changes in satellite glial cells in cultured trigeminal ganglia. Neuron Glia Biol. 2010;6(4):237-43. DOI: 10.1017/S1740925X1100007X
  25. Capuano A, De Corato A, Lisi L, Tringali G, Navarra P, Dello Russo C. Proinflammatory-activated trigeminal satellite cells promote neuronal sensitization: relevance for migraine pathology. Molecular Pain. 2009;5:43.
  26. Locher H, de Rooij KE, de Groot JC, van Doorn R, Gruis NA, Löwik CW, et al. Class III β-tubulin, a novel biomarker in the human melanocyte lineage. Differentiation. 2013;85(4-5):173-81. DOI: 10.1016/j.diff.2013.05.003
  27. Dráberová E, Del Valle L, Gordon J, Marková V, Smejkalová B, Bertrand L, et al. Class III beta-tubulin is constitutively coexpressed with glial fibrillary acidic protein and nestin in midgestational human fetal astrocytes: implications for phenotypic identity. J Neuropathol Exp Neurol. 2008;67(4):341-54. DOI: 10.1097/NEN.0b013e31816a686d
  28. Locher H, Saadah N, de Groot S, de Groot JC, Frijns JH, Huisman MA. Hair follicle bulge cultures yield class III-β-tubulin-positive melanoglial cells. Histochem Cell Biol. 2015;144(1):87-91. DOI: 10.1007/s00418-015-1312-8
  29. Schafer MK, Mahata SK, Stroth N, Eiden LE, Weihe E. Cellular distribution of chromogranin A in excitatory, inhibitory, aminergic and peptidergic neurons of the rodent central nervous system. Regul Pept. 2010;165(1):36-44. DOI: 10.1016/j.regpep.2009.11.021
  30. Xue ZG, Smith J, Le Douarin NM. Expression of the adrenergic phenotype by dorsal root ganglion cells of the quail in culture in vitro. C R Acad Sci III. 1985;300(13):483-8.
  31. Furlan A, Dyachuk V, Kastriti ME, Calvo-Enrique L, Abdo H, Hadjab S, et al. Multipotent peripheral glial cells generate neuroendocrine cells of the adrenal medulla. Science. 2017;357(6346). DOI: 10.1126/science.aal375

Publication of the article:

«Bulletin of problems biology and medicine» Issue 3 (152), 2019 year, 46-50 pages, index UDK 611.“461”