Vasylovska S. Functional characteristics of cord blood progeni-tor cells under in vitro culture conditions

The dissertation is devoted to the investigation of the influence of different culture conditions on the functional activity of cord blood-derived hematopoietic progenitor cells (CB HPC), and determination of the conditions necessary for long-term maintenance of hematopoiesis in vitro. It was shown that the maintenance of hematopoietic activity of CB HPCs in the long term culture can be achieved with the aid of the mitotically inactivated feeder layer derived from the stromal component of the fetal liver (FL) of 7-8 weeks of gestation and from the fetal bone (FB) of the 10-11 weeks of gestation. Maximal increase in the number of nucleated cells and clonogenic activity of CB HPCs was observed on the 3rd week of culture. It was established that the FL feeder layer is more efficient than the FB feeder layer in maintenance of hematopoiesis in vitro. Addition of the exogenous cytokines such as GM-CSF, IL-3, IL-6 and SCF to the culture system containing the FL feeder layer stabilized growth dynamics of the nucleated cells and clonogenic progenitors (CFU-GM) during the 5-week culture, inducing up to an 80 fold increase in the total number of clonogenic progenitors in comparison to the initial suspension. The most pronounced stimulating effect on the AC133+ cells was observed after 16-hour incubation with the Flt-3 growth factor prior to culture. Subsequent long term culture of the AC133+ cells on the FL feeder layer with cytokines resulted in a 55 fold increase in the number of nucleated cells, and an increase of clonogenic progenitors in the fraction from 4,2% to 35,1% on the 5th week of cultivation.