Using 2H2O to study the influence of feeding on protein synthesis: Effect of isotope equilibration in vivo vs. in cell culture

Danielle A. Dufner, Ilya R. Bederman, Daniel Z. Brunengraber, Nadia Rachdaoui, Faramarz Ismail-Beigi, Brett A. Siegfried, Scot R. Kimball, Stephen F. Previs

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Abstract

We previously reported that 2H2O) can be used to measure rates of protein synthesis during prolonged steady-state conditions (Previs SF, Fatica R, Chandramouli V, Alexander JC, Brunengraber H, and Landau BR. Am J Physiol Endocrinol Metab 286: E665-E672, 2004). The underlying premise of our method is that following the administration of 2H 2O, 2H atoms in body water rapidly equilibrate with free alanine before it is incorporated into newly synthesized proteins. We have now directly examined whether 2H2O can be used to measure the influence of a single meal on protein synthesis. In addition, we have compared the use of 2H2O for measuring rates of protein synthesis in vivo vs. in cell culture. Using a rat model, we observed rapid equilibration between 2H in body water and free alanine; therefore we were able to study the response of protein synthesis to a single meal. We observed that ∼50% of the plasma albumin that is synthesized over the course of 24 h is made within ∼5 h after eating (in rats trained to eat a complete 24-h ration of food in a single meal). Contrary to what we observed in vivo, feeding (the replenishment of cell culture medium) does influence the use of 2H2O for in vitro studies. In particular, since there can be slow equilibration of 2H between water and alanine in the cell culture medium, special consideration must be made to avoid under-estimating the rate of protein synthesis in vitro.

Original languageEnglish (US)
Pages (from-to)E1277-E1283
JournalAmerican Journal of Physiology - Endocrinology and Metabolism
Volume288
Issue number6 51-6
DOIs
StatePublished - Jun 1 2005

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All Science Journal Classification (ASJC) codes

  • Endocrinology, Diabetes and Metabolism
  • Physiology
  • Physiology (medical)

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