NMR studies of erythrocytes immobilized in agarose and alginate gels.
31P and 13C NMR were used to study the energy metabolism in perfused, human erythrocytes. The erythrocytes were immobilized in agarose threads, Ca- or Ba-alginate beads, and Ba-alginate-coated agarose threads. Erythrocytes were easily washed out from the agarose threads, but not from alginate-containing gels. Various small molecules, such as hypophosphite, dimethyl methylphosphonate, and methylphosphonate, were taken up from the perfusion medium in a normal manner. In addition, the 2,3-bisphosphoglycerate (2,3-DPG) chemical shifts were sensitive to the oxygen partial pressure suggesting that O2 molecules were diffusing through the gel and modifying the binding of 2,3-DPG to hemoglobin. A combination of inosine and pyruvate stimulated the synthesis of 2,3-DPG, but only if inorganic phosphate was present in the perfusion medium. Inosine only resulted in a dramatic rise in the intracellular sugarphosphate concentrations. Furthermore, [2-13C]glucose was converted to [2-13C]lactate by immobilized cells at a rate which was comparable to that in a control suspension. In summary, immobilization in Ba-alginate-coated agarose threads was an efficient way of trapping human erythrocytes for whole cell NMR investigations.
Lundberg
P
P
Department of Biochemistry, University of Sydney, New South Wales, Australia.
Berners-Price
S J
SJ
Roy
S
S
Kuchel
P W
PW
D000464:Alginates; D004163:Diphosphoglyceric Acids / Q000378:metabolism*; D004734:Energy Metabolism / Q000187:drug effects / Q000502:physiology*; D004912:Erythrocytes / Q000378:metabolism*; D005782:Gels; D006801:Humans; D066298:In Vitro Techniques; D007288:Inosine / Q000494:pharmacology; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D010100:Oxygen / Q000502:physiology; D010313:Partial Pressure; D011773:Pyruvates / Q000494:pharmacology; D012685:Sepharose
D000464:Alginates; D004163:Diphosphoglyceric Acids; D005782:Gels; D011773:Pyruvates; D007288:Inosine; D012685:Sepharose; D010100:Oxygen