DETERMINATION OF SPECIFIC OXYGEN-UPTAKE RATES IN HUMAN HEMATOPOIETIC CULTURES AND IMPLICATIONS FOR BIOREACTOR DESIGN

Oxygen plays an important role in the cultivation of primary cells ex vivo. In this study, we used hermetically sealed tissue culture well i nserts equipped with oxygen electrodes to measure the oxygen utilizati on of cultured human bone marrow mononuclear cells (BM MNCs). The oxyg en uptake rate (OUR) of BM MNCs was determined during a 14-day culture in which both adherent and nonadherent cells were present. Early in t he culture, the cells exhibited very low OURs. The specific OURs (upta ke rate per cell) were at approximately 0.005 mu mol/10(6) cells/hr sh ortly after the initiation of culture. The OUR then increased as the c ultures developed. After about 8 to 10 days of cultivation the specifi c OURs had increased to 0.038 +/- 0.006 and 0.025 +/- 0.005 mu mol/10( 6) cells/hr for adherent and nonadherent cells, respectively, after wh ich no further increase was observed. Based on these oxygen uptake rat e data, a mathematical model of oxygen diffusion was formulated and us e to investigate issues associated with hematopoietic bioreactor desig n, including initial cell density, medium depth, reactor configuration , and oxygen partial pressure. In situ OUR measurements confirmed pred icted oxygen limitations based on the mathematical model and the exper imentally determined OURs. High-density hematopoietic cultures present design challenges in terms of sufficient and uniform delivery of oxyg en to an active hematopoietic culture. These challenges can be met by using parallel-plate bioreactors with thin liquid layers.