Выделение и биологические характеристики стволовых клеток плаценты

Abstract

Objective. Isolation of stem cells (SCs) from different parts of the placenta and description of their biological characteristics. Materials and methods. The placenta was obtained from parturient women aged 22 to 39 years (mean age was 29.0 ± 3.7 years) with a normal singleton pregnancy resulting in normal delivery and with written informed consent obtained at 37–41 weeks of gestation after non-invasive (n = 32) or operative (n = 2) delivery. Wharton’s jelly from the umbilical cord and the fetal side of the placenta, the chorionic plate with chorion frondosum, were isolated and cultured to obtain mesenchymal stem cells (MSCs). Morphological evaluation of cell culture, differentiation, immunological characterization (CD34, CD45, CD90, CD105, CD73, HLA-DR (BD Biosciences and Becman Coulter, USA)), assessment of viability using 7-ADD dye on the FACSCanto II flow cytometry facility (Becton Dickinson CA, USA), cell growth, sterility control, analysis of STR (Short tandem repeat) loci polymorphism for identification, karyotyping, and cryopreservation of cell cultures were carried out. Results. Primary cell cultures were isolated from the chorionic plate with chorion frondosum and Wharton’s jelly of the umbilical cord in 100% of cases. MSCs had high proliferative activity and viability (98.0 ± 1.2% 7-ADD) throughout the whole period of cultivation. The growth rate of MSCs from the chorionic plate at P3, P4, and P5 was higher compared to that of MSCs from the Wharton’s jelly during cultivation, p ≤ 0.05. Immunophenotype corresponded to the requirements of International Society for Cellular Therapy: high expression of MSC markers (CD73, CD90, CD105) was detected; hematopoietic and lymphocytic markers were absent (CD34, CD45, HLA-DR). The potential of cells to differentiate in the mesenchymal direction (chondrocytes, osteocytes, adipocytes) was confirmed in all obtained MSCs. In 95% of cases, the cells were of fetal origin. The karyotype of the examined MSC cell lines was normal: 46, XX (female) or 46, XY (male). All cell cultures were tested, cryopreserved, and placed in a biobank. Conclusion. Isolation and evaluation of biological characteristics of human placental stem cells is of great interest and is a promising direction in regenerative medicine due to the simplicity of placenta sampling, the absence of ethical problems, the ability to quickly obtain and accumulate the necessary amount of cell material with a stable genotype and given biological characteristics. Key words: mesenchymal stem cells, placenta, immunophenotype, STR loci polymorphisms, karyotype