Abstract
Hepatocytes obtained from rats fed a choline-deficient diet for 3 days were cultured in a medium +/- choline (100 microM) or methionine (200 microM). We investigated how choline deficiency affected hepatic lipogenesis, apolipoprotein synthesis, and lipoprotein secretion. The mass of triacylglycerol and phosphatidylcholine secreted was increased about 3-fold and 2-fold, respectively, by the addition of either choline or methionine to the cultured cells. Similarly, a 3-fold stimulation in the secretion of [3H]triacylglycerol and [3H]phosphatidylcholine derived from [3H]oleate was observed after the addition of choline or methionine. Fractionation of secreted lipoproteins by ultracentrifugation revealed that the reduced secretion of triacylglycerol and phosphatidylcholine from choline-deficient cells was mainly due to impaired secretion of very low density lipoproteins (VLDL) (but not high density lipoproteins (HDL)). Fluorography of L-[4,5-3H]leucine-labeled lipoproteins showed a remarkable inhibition of VLDL secretion by choline deficiency. The addition of choline or methionine stimulated the synthesis of phosphatidylcholine and increased the cellular phosphatidylcholine levels to that in normal cells. While there was little effect of choline on the synthesis and amount of cellular phosphatidylethanolamine, the addition of methionine diminished cellular phosphatidylethanolamine levels. Choline deficiency did not change the rate of incorporation of L-[4,5-3H]leucine into cellular VLDL apolipoproteins, nor the rate of disappearance of radioactivity from L-[4,5-3H]leucine-labeled cellular apoB, apoE, and apoC. These results suggest that hepatic secretion of VLDL, but not HDL, requires active phosphatidylcholine biosynthesis. Secondly, the inhibitory effect of choline deficiency on VLDL secretion can be compensated by the methylation of phosphatidylethanolamine.
Highlights
Hepatocytes obtained from rats fed a choline-defi- outside coatof the maturelipoprotein particles contains phoscientdietfor 3 days were cultured in a medium f pholipid, of which PC accounts for 60% of the phospholipid choline (100 p ~ or) methionine (200 p ~ ) W. e investigated how choline deficiency affected hepatic lipogenesis, apolipoprotein synthesis, and lipoprotein secretion
Fractionation of secreted lipoproteins by ultracentrifugation revealed that thereduced secretion of triacylglycerol and phosphatidylcholinefromcholine-deficient cells was mainly due to impaired secretion of methionine[3].Theresults from thislaboratory usinga methylation inhibitor, 3-deazaadenosine, suggested that the methylation pathwaywas not required for hepatic lipoprotein secretion [5]
The rates of incorporation of [3H]leucine into cellular apoBH, apoBL,apoE, and apoC were independent of the addition of choline or methionine (Fig. 7 ), suggesting that there was no effect of the deficiency onapolipoprotein synthesis
Summary
Hepatocytes obtained from rats fed a choline-defi- outside coatof the maturelipoprotein particles contains phoscientdietfor 3 days were cultured in a medium f pholipid, of which PC accounts for 60% of the phospholipid choline (100 p ~ or) methionine (200 p ~ ) W. e investigated how choline deficiency affected hepatic lipogenesis, apolipoprotein synthesis, and lipoprotein secretion. Secretion of VLDL, but not HDL, from cultured rat hepatocytes, and this can be rapidlyreversed by the addition of either choline or methionine.
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