title: Effect of oxygen concentration on viability and metabolism in a fluidized-bed bioartificial liver using ³¹P and ¹³C NMR spectroscopy.
abstract: Many oxygen mass-transfer modeling studies have been performed for various bioartificial liver (BAL) encapsulation types; yet, to our knowledge, there is no experimental study that directly and noninvasively measures viability and metabolism as a function of time and oxygen concentration. We report the effect of oxygen concentration on viability and metabolism in a fluidized-bed NMR-compatible BAL using in vivo ³¹P and ¹³C NMR spectroscopy, respectively, by monitoring nucleotide triphosphate (NTP) and ¹³C-labeled nutrient metabolites, respectively. Fluidized-bed bioreactors eliminate the potential channeling that occurs with packed-bed bioreactors and serve as an ideal experimental model for homogeneous oxygen distribution. Hepatocytes were electrostatically encapsulated in alginate (avg. diameter, 500 μm; 3.5×10⁷ cells/mL) and perfused at 3 mL/min in a 9-cm (inner diameter) cylindrical glass NMR tube. Four oxygen treatments were tested and validated by an in-line oxygen electrode: (1) 95:5 oxygen:carbon dioxide (carbogen), (2) 75:20:5 nitrogen:oxygen:carbon dioxide, (3) 60:35:5 nitrogen:oxygen:carbon dioxide, and (4) 45:50:5 nitrogen:oxygen:carbon dioxide. With 20% oxygen, β-NTP steadily decreased until it was no longer detected at 11 h. The 35%, 50%, and 95% oxygen treatments resulted in steady β-NTP levels throughout the 28-h experimental period. For the 50% and 95% oxygen treatment, a ¹³C NMR time course (∼5 h) revealed 2-¹³C-glycine and 2-¹³C-glucose to be incorporated into [2-¹³C-glycyl]glutathione (GSH) and 2-¹³C-lactate, respectively, with 95% having a lower rate of lactate formation. ³¹P and ¹³C NMR spectroscopy is a noninvasive method for determining viability and metabolic rates. Modifying tissue-engineered devices to be NMR compatible is a relatively easy and inexpensive process depending on the bioreactor shape.
authors: Department of Biomedical Engineering, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
authors: Rex E
authors: RE
authors: Jeffries
authors: Michael P
authors: MP
authors: Gamcsik
authors: Kayvan R
authors: KR
authors: Keshari
authors: Peter
authors: P
authors: Pediaditakis
authors: Andrey P
authors: AP
authors: Tikunov
authors: Gregory B
authors: GB
authors: Young
authors: Haakil
authors: H
authors: Lee
authors: Paul B
authors: PB
authors: Watkins
authors: Jeffrey M
authors: JM
authors: Macdonald
mesh_terms: D000818:Animals; D001187:Artificial Organs; D019149:Bioreactors; D002247:Carbon Isotopes; D008099:Liver / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D010100:Oxygen / Q000378:metabolism*; D010759:Phosphorus Isotopes; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D002247:Carbon Isotopes; D010759:Phosphorus Isotopes; D010100:Oxygen