Cholestane-3β,5α,6β-triol stimulates phospholipid synthesis and CTP-phosphocholine cytidyltransferase in cultured LLC-PK cells

Abstract

The present study was conducted to examine the effect, if any, of triol on the rate of total or individual phospholipid synthesis by LLC-PK cells in culture. LLC-PK cells were incubated in medium with or without 10 μg/ml of 5α-cholestane-3β,5α,6β-triol (triol) for 24 h. Triol-treated and control cells were then incubated with medium containing either [ 14C]glycerol or [ 32P]phosphate for 1, 6 or 12 hr. In triol-treated cells, the amount of labeled glycerol and [ 32P]phosphate incorporated into glycerophospholipids and phospholipids (PL), respectively, were higher in triol-treated cells than in control cells, indicating a higher rate of PL synthesis in triol-treated cells. The results also showed that the increase in PL synthesis was higher in magnitude for some PL than others, thus disturbing the ratios among the PL fractions in the cell membrane. CTP-phosphocholine cytidyltransferase activity was greatly enhanced in the cytosolic as well as the particulate fractions of the triol-treated cells, which explains the increase of PC synthesis under triol effect. The rate of [ 3H]acetate incorporation into the total and free fatty acid fractions was significantly increased in triol-treated cells. The activation of the cytidyl transferase enzyme was related to the enhanced de novo synthesis and cellular uptake of fatty acids in triol-treated cells, which make fatty acids more available in these cells and can upregulate the enzyme. The increased synthesis of phospholipids in the triol cells and the increased level of phopholipid in these cells (as μg lipid phosphorus/mg cell protein) observed in our previous study indicate changes in the phospholipid head group composition of the triol cells. These changes can affect several membrane properties and membrane bound enzymes.