ISSN 2415-3060 (print), ISSN 2522-4972 (online)
  • 7 of 58
УЖМБС 2020, 5(6): 59–65
Experimental Medicine and Morphology

Microscopic Investigation of Compatibility of Samples Containing Multipotent Mesenchimal Stromal Cells of Additive Tissue in Experimental Conditions

Bambuliak A.V., Kuzniak N.B., Dmitrenko R.R., Tkachik S.V., Honcharenko V.A.

The purpose of the study was to investigate the biocompatibility of samples containing multipotent mesenchymal stromal cells of adipose tissue to replace bone defects. Material and methods. The study was conducted at Bukovina State Medical University, Chernivtsi, Ukraine. Adipose tissue samples were obtained from the neck of 60 experimental animals (white Wistar rats). We selected 4 samples for the toxicological experiment, which allowed to establish the direct influence of factors in the contact of implantation material at the cellular level. Sample № 1 - Multipotent mesenchymal stromal cells of adipose tissue, which underwent osteogenic differentiation; № 2 - Multipotent mesenchymal stromal cells of adipose tissue with osteogenic differentiation with the addition of platelet-enriched blood plasma; № 3 - “Kolapan” with applied tissue culture of Multipotent mesenchymal stromal cells of adipose tissue cells, which underwent osteogenic differentiation; № 4 - "Kolapan" + Multipotent mesenchymal stromal cells of adipose tissue + platelet-enriched plasma. Multipotent mesenchymal stromal cells of adipose tissue were obtained by grinding adipose tissue of rats in 0.1% collagenase 1A [14]. The study of biocompatibility by cell culture in vitro was performed in accordance with the Working Instruction № 04/2013-VL. The cultures were investigated by the explantation method in a plasma clot in Karelian vials. In order to standardize the nature of growth, their zones were classified into compact, reticular and migrating cells of growing fibroblastic tissues. To assess the probability of the obtained results of the study we used a variation-statistical method of analysis using Microsoft Excel. Statistical calculation of the results of clinical and laboratory studies was carried out according to conventional methods. Results and discussion. Microscopic examination of the surface of samples with culture of fibroblasts showed their satisfactory adhesion on the tooth surface after 5 days of cultivation. In the study of sample № 1 (Multipotent mesenchymal stromal cells of adipose tissue, with osteogenic differentiation), it was noted that the structure of the cells acquired a rounded and oval shape, which indicated their destruction and damage. On the 5th day of observation, cells with numerical intussusception and processes were observed during visual examination of sample № 3 (“Kolapan”, with applied culture of Multipotent mesenchymal stromal cells of adipose tissue). In the study of samples № 4 (Multipotent mesenchymal stromal cells of adipose tissue + platelet-enriched plasma + "Kolapan") on the 5th day of research, signs of growth were manifested by migration of fibroblastic elements that had a spindle-shaped and polygonal shape, with the formation of the primary zone due to strands. On the 7th day of cultivation in experimental samples № 2, № 3, № 4 there was the formation of three growth zones: compact - from cells of polygonal and spindle-shaped form; reticulate - from strands and bundles of cells that were located reticulate and areas of single migrating elements of spindle-shaped. External characteristics and cell growth surface did not differ from control samples. On the 10th day of cultivation in the experimental samples, as well as in the control, the areas of compact and reticular growth zone and the zone of migrating fibroblasts were increased. At the same time, tissue-like growth of cells was observed. Visualization of compact and stack-like zones of the studied experimental samples revealed signs of the beginning of degenerative changes, which was characterized in the form of rounding of the shape and vacuolation of cells. This trend was most pronounced in samples № 2 and № 4. Conclusion. Thus, tissue equivalents of bone tissue based on Multipotent mesenchymal stromal cells of adipose tissue can be candidates for use in regenerative medicine, and studies of their application in experimental animals will provide an opportunity to expand the understanding of the characteristics of Multipotent mesenchymal stromal cells of adipose tissue in order to optimize their further clinical application and implement new approaches in different areas of dentistry

Keywords: multipotent mesenchymal stromal cells of adipose tissue, platelet-enriched plasma, osteogenic differentiation, growth zones

Full text: PDF (Ukr) 463K

  1. Coleman SR, Mazzola RF, Q L, Lee Pu. Fat Injection: from filling to regeneration. 2nd ed. NY: CRC Press; 2016. 900 p.
  2. Mazurkevych AI, Maliuk MO, Tkachenko SM, Kharkevych, YuO. Study of biocompatibility of hemostatic sponges with thebarrel cages of marrow of rabbit during cultivation of in vitro. Bulletin of the Sumy National Agrarian University. 2015; 1(34): 7-11.
  3. Aggarwal S, Pittenger MF. Human mesenchymal stem cells modulate allogeneic immune cell responses. Blood. 2015; 105: 1815-22. PMID: 15494428.
  4. Augello A, Tasso R, Negrini SM, Amateis A, Indiveri F, Cancedda R, et al. Bone marrow mesenchymal progenitor cells inhibit lymphocyte proliferation by activation of the programmed death 1 pathway. Eur J Immunol. 2019; 43: 1482-90. PMID: 15827960.
  5. Mangashetti LS, Khapli SM, Wani MR. IL-4 inhibits bone-resorbing activity of mature osteoclasts by affecting NF-kappa B and Ca2+ signaling. J Immunol. 2017; 175: 917-25. PMID: 16002690.
  6. Wang Y, Du Y, Yuan H, Pan Y, Wu J, Du X, et al. Human amnion-derived mesenchymal stem cells enhance the osteogenic differentiation of human adipose-derived stem cells by promoting adiponectin excretion via the APPL1-ERK1/2 signaling pathway. IUBMB Life. 2020; 72(2): 296-304.
  7. Roodman GD. Role of cytokines in the regulation of bone resorption. Calcif Tissue Int. 2013; 1(61): 94-8. PMID: 8275387.
  8. Pittenger MF, Mackay AM, Beck SC, Jaiswal RK, Douglas R, Mosca JD, et al. Multilineage potential of adult human mesenchymal stem cells. Science. 2015; 284: 143-7. PMID: 10102814.
  9. Nifant'ev I, Bukharova T, Dyakonov A, Goldshtein D, Galitsyna E, Kosarev M, et al. Osteogenic differentiation of human adipose tissue-derived MSCs by non-toxic calcium poly(ethylene phosphate)s. Int J Mol Sci. 2019; 20(24): 6242.
  10. Yamanaka S. Pluripotency and nuclear reprogramming. Philosophical Transactions of the Royal Society of London. Series B: Biological Sciences. 2016; 363(1500): 2079-87. PMID: 18375377. PMCID: PMC2610180.
  11. Amabile G, Meissner A. Induced pluripotent stem cells: current progress and potential for regenerative medicine. Trends in Molecular Medicine. 2014; 15(2): 59-68. PMID: 19162546.
  12. Friedenstein AJ, Petrakova KV, Kurolesova AI, Frolova GP. Heterotopic of bone marrow.Analysis of precursor cells for osteogenic and hematopoietic tissues. Transplantation. 2014; 6: 230-47. PMID: 5654088.
  13. Baer PC, Koch B, Hickmann E, Schubert R, Cinatl J Jr, Hauser IA, et al. Isolation, characterization, differentiation and immunomodulatory capacity of mesenchymal stromal/stem cells from human perirenal adipose tissue. Cells. 2019; 8(11):0.
  14. Yanai R, Tetsuo F, Ito S, Itsumi M, Yoshizumi J, Maki T, et al. Extracellular calcium stimulates osteogenic differentiation of human adipose-derived stem cells by enhancing bone morphogenetic protein-2 expression. Cell Calcium. 2019; 83: 102058.