Metabolic characterization of T cell lymphoblastoid, Jurkat, in the presence of respiration inhibitors in a continuous reactor


Commercial scale mammalian cell culture systems have many technical barriers: (i) accumulation of inhibitory waste products; (ii) sensitivity to low level of shear; and (iii) cellular oxygen demand in excess of bioreactor's capability to provide oxygen without damaging cells. Because of these problems, systematic approaches for providing sufficient oxygen transfer rates to large scale mammalian cell culture systems have yet to be adequately developed. This work, however, suggests a unique approach to this problem from a different point of view. A research program has been undertaken to determine if the metabolism of mammalian cells can be manipulated to minimize or eliminate the oxygen transfer capabilities required by the cells from a given bioreactor. The respiration inhibitor, rotenone, has been added to the cultures of a T cell fusion partner, Jurkat, at levels that completely inhibit respiration. Growth of the cells, at these concentrations of rotenone in completely defined media, did not occur unless the media was supplemented with at least 1 mM pyruvate or at least 1% fetal bovine serum. Respiration inhibited cells utilized much less glutamine, respiratory fuel, and concomitantly produces much less ammonia. They grew to one million cells per ml at near normal rate but the maximum cell density was 65% that of un-inhibited cultures. Proline and a minimum amount of glutamine (0.2 mM) were found to be essential for the growth of Jurkat cells in the presence of rotenone. Intracellular thiol content measured with flow cytometry technique was found to be lower for rotenone cultures, possibly due to the free radicals formation at higher oxygen concentration. With continuous reactor experiments, the fact that Jurkat cells grow without using oxygen in the presence of rotenone was firmly confirmed and more metabolic details were found. In addition to the increased glycolysis, respiration inhibited culture increased the utilization of arginine, asparagine, serine, histidine, and aspartic acid and increased the production of lysine, glycine, glutamic acid, and alanine. At steady state with 90 nM of rotenone, pulse inputs of proline and pyruvate increased the cell density up to 1.4 million cells per milliliter

Similar works

Full text


DSpace at Rice University

Full text is not available
oaioai:scholarship.rice.e...Last time updated on 6/11/2012

This paper was published in DSpace at Rice University.

Having an issue?

Is data on this page outdated, violates copyrights or anything else? Report the problem now and we will take corresponding actions after reviewing your request.