Using H218O to study protein and lipid flux: Is dyslipidemia a problem of triglyceride and/or apoB production?

Abstract

Investigations surrounding dyslipidemia often examine lipid (e.g. triglyceride) and protein (e.g. apoB) kinetics in separate studies which is unfortunate since the pathways are related. We recently demonstrated the ability to quantify protein flux in vivo using H2 18O; the hydrolysis of pre‐existing proteins in H2 18O generates labeled amino acids, thus protein synthesis is estimated from the incorporation of those labeled amino acids into newly made proteins. We now demonstrate a similar logic for studying lipid synthesis, i.e. the degradation of pre‐existing lipids in H2 18O labels fatty acids, thus lipid synthesis is estimated from the incorporation of 18O‐fatty acids into different end‐products using LC‐MS/MS. Although the chromatographic runs are complex (numerous analytes co‐elute) it is possible to measure the labeling of specific lipids. We demonstrate the application of this method in studies of triglyceride and apoB production; it seems possible to uncouple triglyceride and apoB flux in C57Bl/6J mice under different conditions, e.g. control vs Intralipid or MTPi‐treated. The method described herein is suitable for routine investigations in rodents, the tracer is given via intraperitoneal injection and the analyses require 1 to 2 plasma samples (each ≤40 μL). Our approach should enable “high‐throughput” in vivo studies regarding the pathophysiology of dyslipidemia.