Study On Ex Vivo Expansion Of Human Umbilical Cord Blood Hematopoietic Cells

This paper mainly study on several kind of cytokines, culture system parameters and finally find out the optimum culture parameters. The main result are as follows:(1) When preserved in 4℃ refrigerator,the vitality and concentration of MNC in cord blood diminished gradually, The ability of CFU producing declined rapidly when exceed 2d,In the 5th day the output of CFU dropped to 1/10 of the output of 1th day. In the sixth day , MNC in cord blood lost the ability of producing CFU. When preserved in the IMDM media ,the viality and ability of producing CFU dropped more rapidly ,In the 4 th day of preservation the MNC lost the ability of producing CFU, and in the 5 th day of preservation the viality of MNC dropped to 0. So that ,When preserved in the 4℃ refrigerator, the MNC should preserved in the condition of complete cord blood ,and the period of preservation should not exceed 3 days.(2) The optimum combination of cytokines is :SCF+IL-6+FL+TPO, and the respective optimum concentration of each cytokines is : 50, 20, 80,40ng/ml, The cytokines FL most effectively infience the expansion of CFU-C .,MNC expanded gradually During 1~3 weeks and CFU-C expanded gradually during the 1-2 weeks, When between 2~3weeks ,the trend of the expansion dropped to some level. With the culture period prolonged ,the output of MNC was going up .then the output of CFU of majority of the research groups dropped gradually.So that , Hematopoietic cells should be harvested within 3 weeks of culture.(3) When only add FL to the Medium, MNC and CFU-C can not expanse effectively, and the consumption of FL was not abundantly. When add cytokines combination into the medium ,The expansion of MNC and CFU-C turn remarkable, and the consumption of FL increased rapidly. The FL consumption has some relationship to the CFU-C expansion. Cytokines combination can promote the growth of hematopoietic cells and the consumption of cytokines(4) The optimum culture system parameters are as follows: pH: 7.2 mixed CBP concentration:20% > MNC inoculum:4 X 105/ml . frequency of changing medium:every 3 days. pH and MNC inoculum affected the expansion of hematopoietic cells deepest and concentration of mixed CBP influence the expansion of hamatopoietic cells slightly.