In studies designed to measure rates of hepatic cholesterogenesis, rat liver slices were incubated with [14C] acetate in the presence of unlabeled octanoate and with [14C] octanoate the presence of unlabeled acetate to assess the degree of intracellular dilution and compartmentalization of the acetyl CoA pools. It was observed that when increasing concentrations of [14C] acetate were used, the incorporation rate of this substrate into cholesterol became constant under circumstances where the specific activity of the ketone bodies continued to rise. The addition of unlabeled acetate reduced the flow of radiolabeled C2 units from [14C] octanoate into cholesterol but not into ketone bodies or CO2. Use of the specific activity of the ketone bodies was shown to yield an appropriate correction for the flux of [14C] octanoate into cholesterol: in contrast, this correction was shown to be inappropriate in experiments in which cholesterogenesis was measured using radiolabeled acetate. Taken together, these findings indicate that [14C] acetate gives rise to a cytosolic pool of acetyl CoA that is used for cholesterol synthesis but is not in isotopic equilibrium with the mitochondrial pool of acetyl CoA used for ketogenesis. From a combination of experiments using acetate and octanoate, singly and together and alternately labeled, the apparent and absolute fluxes of C2 units through both pools were measured. Maximum rates of sterol synthesis were obtained in the presence of octanoate. In contrast, with [14C] acetate as substrate the flux of labeled C2 units into cholesterol was only 47% of maximum while total C2 flux was 75% of that observed with octanoate. Thus, in addition to the errors associated with intracellular dilution when [14C] acetate is used, a further limitation to the use of this substrate appears to be the presence of a partially rate limiting step in its metabolism prior to hydroxymethylglutaryl CoA reductase.
|Original language||English (US)|
|Number of pages||7|
|Journal||Journal of Biological Chemistry|
|State||Published - 1974|
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology