The effects of fetal neural cell conditioned medium on cell proliferation in the rat brain after traumatic brain injury

Home/2021, Vol. 9, No. 2/The effects of fetal neural cell conditioned medium on cell proliferation in the rat brain after traumatic brain injury

Cell and Organ Transplantology. 2021; 9(2):in press.
DOI: 10.22494/cot.v9i2.126

The effects of fetal neural cell conditioned medium on cell proliferation in the rat brain after traumatic brain injury

Lisyany M.1, Stanetska D.1, Govbakh I.2, Tsupykov O.3,4

  • 1A. P. Romodanov State Institute of Neurosurgery, National Academy of Medical Sciences of Ukraine, Kyiv, Ukraine
  • 2Kharkiv Medical Academy of Postgraduate Education, Ministry of Public Health of Ukraine, Kharkiv, Ukraine
  • 3Bogomoletz Institute of Physiology, National Academy of Sciences, Kyiv, Ukraine
  • 4State Institute of Genetic and Regenerative Medicine, National Academy of Medical Sciences of Ukraine, Kyiv, Ukraine


Traumatic brain injury (TBI) is accompanied by an increase in the number of proliferating cells. However, the question of the nature, conditions of production and mechanisms of action of humoral factors secreted by fetal neural cells (FNCs) on reparative processes and neurogenesis in the brain after trauma and FNCs transplantation remains open.
The purpose of the study was to establish the possibility of the influence of the conditioned medium of fetal neural cell cultures on the proliferative activity of Ki-67-positive cells in the cortex and subcortical structures of the rat brain after TBI.
Materials and methods. TBI was simulated by dropping a metal cylinder on the rat’s head. Rats (E17-18) were used to obtain cultures of neural stem/progenitor cells. Conditioned media from cell cultures with high adhesive properties (HA-CM) and low adhesive properties (LA-CM) were used to treat the effects of experimental TBI in rats by intramuscular injection. The effect of conditioned media on the proliferative activity of Ki-67-positive cells in the cortex and subcortical structures of the brain after TBI was determined by immunohistochemical analysis using antibodies against Ki-67 protein.
Results. Immunohistochemical analysis of the brain sections showed that on the 5th day after traumatic brain injury in rats there was a probable increase in the number of Ki-67-positive cells in the cortex, hippocampus and thalamus. It was found that the injection of HA-CM or LA-CM in animals with TBI increased the number of Ki-67-positive cells in the hippocampus compared with the TBI group and their value for the TBI+LA-CM group reached 59.6 ± 6.1, and for the TBI+HA-CM group – 47.2 ± 3.1 cells (p <0.05 compared with the TBI group). In the cortex and thalamus, the number of Ki-67-positive cells in contrast decreased compared with the group of animals with TBI and for the group TBI+LA-CM was 20.2 ± 1.6 and 12.0 ± 1.7, respectively, and for the group TBI+HA-CM – 25.3 ± 2.1 and 13.3 ± 1.3, respectively.
Conclusions. The administration of LA-CM or HA-CM to animals with traumatic brain injury increases the number of Ki-67-positive cells in the hippocampus, possibly associated with increased neurogenesis, and decreases in the cortex and thalamus, which may be due to a weakening of reactive gliosis.

Key words: traumatic brain injury; fetal neural cells; conditioned medium; immunohistochemistry; proliferative activity


1. Sabet N, Soltani Z, Khaksari M. Multipotential and systemic effects of traumatic brain injury. J Neuroimmunol. 2021; 357:577619. DOI: 10.1016/j.jneuroim.2021.577619.
2. Neuberger EJ, Swietek B, Corrubia L, Prasanna A, Santhakumar V. Enhanced dentate neurogenesis after brain injury undermines long-term neurogenic potential and promotes seizure susceptibility. Stem Cell Reports. 2017; 9(3):972-84. DOI: 10.1016/j.stemcr.2017.07.015.
PMid:28826852 PMCid:PMC5599224
3. Rizk M, Vu J, Zhang Z. Impact of pediatric traumatic brain injury on hippocampal neurogenesis. Neural Regen Res. 2021; 16(5):926-33. DOI: 10.4103/1673-5374.297057.
PMid:33229731 PMCid:PMC8178782
4. Chen XH, Iwata A, Nonaka M, Browne KD, Smith DH. Neurogenesis and glial proliferation persist for at least one year in the subventricular zone following brain trauma in rats. J Neurotrauma. 2003; 20(7):623-31. DOI: 10.1089/089771503322144545.
5. Jacob B, Osato M. Stem cell exhaustion and leukemogenesis. J Cell Biochem. 2009; 107(3):393-9. DOI: 10.1002/jcb.22150.
6. Encinas JM, Michurina TV, Peunova N, Park JH, Tordo J, Peterson DA, et al. Division-coupled astrocytic differentiation and age-related depletion of neural stem cells in the adult hippocampus. Cell Stem Cell. 2011; 8(5):566-79. DOI: 10.1016/j.stem.2011.03.010.
PMid:21549330 PMCid:PMC3286186
7. Sun D. The potential of endogenous neurogenesis for brain repair and regeneration following traumatic brain injury. Neural Regen Res. 2014; 9(7):688-92. DOI: 10.4103/1673-5374.131567.
PMid:25206873 PMCid:PMC4146269
8. Villasana LE, Kim KN, Westbrook GL, Schnell E. Functional integration of adult-born hippocampal neurons after traumatic brain injury (1,2,3). eNeuro. 2015; 2(5):ENEURO.0056-15.2015. DOI: 10.1523/ENEURO.0056-15.2015.
PMid:26478908 PMCid:PMC4603252
9. Kernie SG, Parent JM. Forebrain neurogenesis after focal ischemic and traumatic brain injury. Neurobiol Dis. 2010; 37(2):267-74. DOI: 10.1016/j.nbd.2009.11.002.
PMid:19909815 PMCid:PMC2864918
10. Sun D, Daniels TE, Rolfe A, Waters M, Hamm R. Inhibition of injury-induced cell proliferation in the dentate gyrus of the hippocampus impairs spontaneous cognitive recovery after traumatic brain injury. J Neurotrauma. 2015; 32(7):495-505. DOI: 10.1089/neu.2014.3545.
PMid:25242459 PMCid:PMC4376486
11. Neuberger EJ, Wahab RA, Jayakumar A, Pfister BJ, Santhakumar V. Distinct effect of impact rise times on immediate and early neuropathology after brain injury in juvenile rats. J Neurosci Res. 2014; 92(10):1350-61. DOI: 10.1002/jnr.23401.
PMid:24799156 PMCid:PMC4300992
12. Schepici G, Silvestro S, Bramanti P, Mazzon E. Traumatic Brain Injury and Stem Cells: An Overview of Clinical Trials, the Current Treatments and Future Therapeutic Approaches. Medicina (Kaunas) 2020; 56(3):137.
PMid:32204311 PMCid:PMC7143935
13. Zhang Y, Chopp M, Zhang ZG, Katakowski M, Xin H, Qu C, et al. Systemic administration of cell-free exosomes generated by human bone marrow derived mesenchymal stem cells cultured under 2D and 3D conditions improves functional recovery in rats after traumatic brain injury. Neurochem Int. 2017; 111:69-81. DOI: 10.1016/j.neuint.2016.08.003
PMid:27539657 PMCid:PMC5311054
14. Chen X, Katakowski M, Li Y, Lu D, Wang L, Zhang L, et al. Human bone marrow stromal cell cultures conditioned by traumatic brain tissue extracts: growth factor production. J Neurosci Res. 2002; 69(5):687-91. DOI: 10.1002/jnr.10334.
15. Shahror RA, Linares GR, Wang Y, Hsueh SC, Wu CC, Chuang DM, et al. Transplantation of mesenchymal stem cells overexpressing fibroblast growth factor 21 facilitates cognitive recovery and enhances neurogenesis in a mouse model of traumatic brain injury. J Neurotrauma. 2020; 37(1):14-26. DOI: 10.1089/neu.2019.6422.
PMid:31298621 PMCid:PMC6921331
16. Bonilla C, Zurita M. Cell-based therapies for traumatic brain injury: therapeutic treatments and clinical trials. Biomedicines. 2021; 9(6):669. DOI: 10.3390/biomedicines9060669
PMid:34200905 PMCid:PMC8230536
17. Olanow CW, Goetz CG, Kordower JH, Stoessl AJ, Sossi V, Brin MF, et al. A double-blind controlled trial of bilateral fetal nigral transplantation in Parkinson’s disease. Ann Neurol. 2003; 54:403-414. DOI: 10.1002/ana.10720.
18. Furtado S, Sossi V, Hauser RA, Samii A, Schulzer M, Murphy CB, et al. Positron emission tomography after fetal transplantation in Huntington’s disease. Ann Neurol. 2005; 58:331-337. DOI: 10.1002/ana.20564.
19. Romanova GA, Shakova FM, Parfenov AL. Modelirovanie cherepno-mozgovoy travmy [Simulation of traumatic brain injury]. Pat fiz eksper ter – Pathophys Fiziol Exp Ther. 2015; 59(2):112-5. [in Russian]
20. Biloshytsky VV. Printsipy modelirovaniya cherepno-mozgovoy travmy v eksperimente [Principles of modeling traumatic brain injury in experiment]. Ukraïnsʹkij nejrohìrurgìčnij žurnal[ – Ukr Neurosurg J. 2008; 4:9-15. [in Russian]
21. Zozulya YuA, Lisyaniy NI. Neyrogennaya differentsirovka stvolovikh kletok [Neurogenic differentiation of stem cells]. Кiev, 2005. 363 p. [in Russian]
22. Kochetov GA. Prakticheskoe rukovodstvo po enzimologii. Metody opredeleniya belka [A practical guide to enzymology. Protein determination methods]. Moscow, 1980. 272 p. [in Russian]
23. Clark LR, Yun S, Acquah NK, Kumar PL, Metheny HE, Paixao RCC, et al. Mild traumatic brain injury induces transient, sequential increases in proliferation, neuroblasts/immature neurons, and cell survival: a time course study in the male mouse dentate gyrus. Front Neurosci. 2021; 14:612749. DOI: 10.3389/fnins.2020.612749.
PMid:33488351 PMCid:PMC7817782
24. Boghdadi AG, Teo L, Bourne JA. The neuroprotective role of reactive astrocytes after central nervous system injury. J Neurotrauma. 2020; 37(5):681-91. DOI: 10.1089/neu.2019.6938.23.
25. Remnant L, Kochanova NY, Reid C, Cisneros-Soberanis F, Earnshaw WC. The intrinsically disorderly story of Ki-67. Open biology. 2021; 11(8):210120. DOI: 10.1098/rsob.210120.
26. Červenka J, Tylečková J, Kupcová Skalníková H, Vodičková Kepková K, Poliakh I, Valeková I, et al. Proteomic characterization of human neural stem cells and their secretome during in vitro differentiation. Front Cell Neurosci. 2021; 14:612560. DOI: 10.3389/fncel.2020.612560
PMid:33584205 PMCid:PMC7876319
27. Einstein O, Friedman-Levi Y, Grigoriadis N, Ben-Hur T. Transplanted neural precursors enhance host brain-derived myelin regeneration. J Neurosci. 2009; 29(50):15694-15702. DOI: 10.1523/JNEUROSCI.3364-09.2009
PMid:20016084 PMCid:PMC6666185

Lisyany M, Stanetska D, Govbakh I, Tsupykov O. The effects of fetal neural cell conditioned medium on cell proliferation in the rat brain after traumatic brain injury. Cell Organ Transpl. 2021; 9(2):in press. doi:10.22494/cot.v9i2.126

Creative Commons License
Is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.