Lysosomal phospholipids from rat liver after treatment with different drugs.

Rats were treated with 5 different drugs p-ethoxyacetanilide (I), indometacin (II) and nor-amidopyrine-methanesulfonate (III), O,O'-bis(diethylaminoethyl)hexestrol(IV) and choloroquine (V) for 3 - 4 weeks. Liver cell fractions were isolated by discontinuous gradient centrifugation and the specific activity of acid phosphatase was determined in each. Lysosomal fractions contained widely varying amounts of this marker enzyme, indicating that the concentration of lysosomes within these fractions differed. The amounts and patterns of phospholipids reflected this fact. Since we assumed bis(monoacylglycero)phosphate [(MAG)2-P; synonym:lysobisphosphatidic acid] is a marker lipid for secondary lysosomes, we expected and found significant quantities of this acidic phospholipid only in those lysosomal fractions which were also rich in acid phosphatase activity. 12% of the lysosomal phospholipids from animals receiving the hexestrol derivative (IV), and 19% of those from the chloroquine (V) experiment were present as (MAG)2P. The fatty acid compositions of this lysosomal phospholipid were not the same in all lysosome fractions. The more (MAG)2P present in the lysosomes, the more unsaturated are the fatty acids. Thus, after treatment with chloroquine, more than 90% of the fatty acids from (MAG)2P are unsaturated; C22:6 represents about 70% of the total.

Lysosomal bis (monoacylglycero)phosphate of rat liver, its induction by chloroquine and its structure.

Since chloroquine is able to induce phospholipidosis, and in particular formation of bis(monoacylglycero)phosphate [(MAG)2P] in rat liver lysosomes, we tested the influence of the drug on this acidic phospholipid 1) after application of the drug for different periods, 3, 7, 14, 21 and 28 days, and 2) after application of different doses of the drug for 7 days. In both cases, the amounts of the total phospholipids and of (MAG)2P within the lysosomal lipids showed a direct relationship to the quantity of the drug. Lysosomal fractions were identified by acid phosphatase activity and by electron microscopy. After 14 days of treatment, the lysosomal fractions showed an increase in the marker enzyme to more than 30 times the level in the homogenate, and consisted of almost pure "lamellar bodies". (MAG)2P was isolated from those lysosomal lipid fractions by column chromatography. We also isolated this phospholipid from "tritosomal" lipids and tested the two preparations in different ways. They both showed glycerophosphoglycerol as the only P-containing product after mild alkaline hydrolysis, and molar ratios for phosphorus: fatty acid: glycerol close to 1:2:2; and yielded diacetylacyl glycerol after acetolysis. Thus, there is no doubt that (MAG)2P occurs in these secondary lysosomes--as we described in others--and that its production is a consequence of accumulation of lysosomes. There is some indication for the occurrence of an acylated (MAG)2P (which is the same as acylphosphatidylglycerol) within the same cell organelles.

On the biosynthesis of plasmalogens during myelination in the rat. VIII[1]. Incorporation of 1-[1-14C] alkyl-2-acyl-3-sn-glycerophosphoethanolamine with different fatty acids.

The uptake of 14C labelled alkyl chains into the alkenyl chains of plasmalogens was estimated 12, 24, 48 and 72 h after intracerebral administration of different substrates to 14 day-old rats We used as substrates 1-[1-14C] alkyl-3-sn-glycerophosphoethanolamines, containing in the 2-position of the glycerol residue a stearoyl (substrate I), an oleoyl (II), an archidonoyl (III), a 4, 7, 10, 13, 16, 19-docosahexaenoyl (IV) or no acyl residue (substrate V, so-called lysoether phosphatide). 1) If the fatty acid in the 2-position of the substrate is a saturated one (as in experiment I), the recovery of radioactive alkyl chains is relatively high, but their desaturation is very slow and seems not to reach its maximum even after 72 h. 2) If a substrate with an unsaturated fatty acid in the 2-position is applied, the number of radioactive chains (alkyl plus alkenyl) depends on the chain length and/or the number of double bonds in the fatty acids. The desaturation of the alkyl chains, however, reaches its maximum about 48 h after application (except for substrate IV). 3) Not only is the recovery of radioactivity in the glycerophosphoethanolamine lipids by far the highest in experiment V at any time, but also the total activity of the alkenyl chains has already reached a maximum 24 h after application.