CULTIVATION AND CHARACTERISTICS OF MESENCHyMAL STEM CELLS FROM THE BONE MARROW OF THE PATIENTS WITH ORTHOPEDIC PATHOLOGY
Nataliya Mikhaylovna ASTAKHOVA1,2, Anastasiya Viktorovna KOREL1, Konstantin Evgen’evich ORISHCHENKO3, Yaroslav Reyngol’dovich EFREMOV3,4, Gleb Aleksandrovich KUDROV KUDROV1, Irina Anatol’evna KIRILOVA1
1Novosibirsk Research Institute of Traumatology and Orthopaedics n.a. Ya.L. Tsivyan of Minzdrav of Russia
2Innovative Medical Technology Center (Medical TechnoPark)
3Institute of Cytology and Genetics of SB RAS
4Novosibirsk State University
Keywords: мезенхимальные стромальные клетки, костный мозг, диспластический коксоартроз, митотический индекс, скорость миграции, дифференцировка, регенерация кости и хряща
Abstract
The article presents data on the cultivation and characteristics of mesenchymal stem cells (MSC) isolated from the bone marrow of patients with dysplastic coxarthrosis. Several morphological phenotypes were found in the fraction of adhesive MSC: spindle-shaped elongated cells, large flattened cells, and thin stellate cells in both samples of bone marrow. Immunophenotypic analysis showed that cells express surface antigens (CD90, CD73, CD105, CD45 and CD34), which are characteristic for typical stem cells. It was shown that the use of a new growth medium containing no components of animal origin for the cultivation of human MSC allowed to achieve confluence of the cell culture on the 16th-8th day of incubation without delaying the proliferative activity of the cells and without loss of ability to differentiate into chondro- and osteogenic types of tissues. Multipotency of MSC was confirmed by osteogenic and chondrogenic differentiation of cells, during prolonged cultivation of MSCs in induction media in vitro . The differentiation of MSC into osteoblasts was confirmed by immunocytochemical staining for alkaline phosphatase and alizarin red S. Specific differentiation of MSC in chondrogenic type was revealed by staining of cartilage deposits with alcian blue. For the first time, such characteristics of human MSC as: mitotic index, trajectory of cells migration and average speed of migration on culture plastics were determined. The mitotic index of actively proliferating MSC was from 2.7 to 3.4 % of the total cell number. The moving activity (speed of cell migration) was 38-42 μm/h. Thus, bone marrow aspirate from patients with orthopedic pathology is the source of stem cells that meet all the criteria for MSC as determined by the International Society of Cellular Therapy and can be used in regenerative therapy of bone and cartilage.
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