Attenuation of Slc27a5 gene expression followed by LC-MS measurement of bile acid reconjugation using metabolomics and a stable isotope tracer strategy

The purpose of this study was to evaluate the use of high resolution LC-MS together with metabolomics and D(4)-cholic acid (D(4)-CA) as a metabolic tracer to measure the metabolism and reconjugation of bile acids (BAs) in vitro and in vivo. Metabolic tracers are very important because they allow for the direct detection (substrate-to-product) of small and significant biological perturbations that may not be apparent when monitoring "static" endogenous levels of particular metabolites. Slc27a5, also known as fatty acid transport protein 5 (FATP5), is the hepatic BA-CoA ligase involved in reconjugating BAs during enterohepatic BA recycling. Using Slc27a5-cKD mice, silencing of ∼90% gene expression was achieved followed by reduction in the reconjugation of D(4)-CA to D(4)-taurocholic acid (D(4)-TCA), as well as other conjugated BA metabolites in plasma (p = 0.0031). The method described allowed a rapid measure of many D(4) and endogenous BA. Analysis of bile resulted in the detection of 39 BA metabolites from a 13 min analytical run. Finally, the utilization of a novel high resolution mass spectrometry method in combination with metabolomics and a stable isotope metabolic tracer allowed for the detection of targeted and untargeted BAs following silencing of the Slc27a5 gene in primary hepatocytes and in mice.


J.M. Castro-Perez, T.P. Roddy, V. Shah, S.P. Wang, X.S. Ouyang, A. Ogawa, D.G. McLaren, M. Tadin-Strapps, M.J. Robinson, S.R. Bartz, B. Ason, Y. Chen, S.F. Previs, K.K. Wong, R.J. Vreeken, D.G. Johns, B.K. Hubbard, T. Hankemeier, L. Mitnaul
Authors from the NMC: 
2011; 10 (10): 4683-4691
Published in: 
Journal of Proteome Research
Date of publication: 
October, 2011
Status of the publication: