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This study was designed to identify a method for the measurement of human high density lipoprotein subfraction (HDL2 and HDL3) metabolism. Apolipoproteins A-I, A-II, and C, the major HDL apoproteins, were radioiodinated and incorporated individually into HDL2 and HDL3 in vitro. Using a double label technique, the turnover of apoA-I in HDL2 and HDL3 was measured simultaneously in a normal male. The apoprotein exchanged rapidly between the two subfractions, evidenced by equilibration of their apoA-I specific activity. Radiolabeled apoA-II, incorporated into the subfractions, showed a similar exchange in vitro. Incubation of 131I-labeled very low density lipoproteins (VLDL) with HDL or its subfractions resulted in transfer of C proteins from VLDL to the HDL moiety. The extent of transfer was dependent on the HDL subfraction present; 50% of the VLDL apoC was transferred to HDL3, while the transfer to total HDL and HDL2 was 69% and 78%, respectively. ApoC also exchanged between HDL2 and HDL3, again showing a preference for the former and suggesting a primary metabolic relationship between VLDL and HDL2. Overall, the study indicates that apoA-I, apoA-II, and the C proteins exist in equilibrium between HDL2 and HDL3. This phenomenon precludes their use as probes for HDL subfraction metabolism in humans.

High density lipoprotein subfractions 2 and 3 were isolated from the sera of normolipidaemic males by rate zonal ultracentrifugation. The fatty acyl compositions of their major lipid classes--phospholipids, cholesteryl esters and triacylglycerols--were determined. The average chain length and degree of unsaturation of the fatty acyl components of the phospholipids and cholesteryl esters of the HDL3 subfraction were found to be significantly higher than those of the HDL2 subfraction. The triacylglycerol moieties of the HDL3 subfraction had a higher proportion of unsaturated fatty acyl components than HDL2, but there was no difference in the average chain length between the two subfractions. These results are not consistent with an equilibration of the lipid components between the two HDL subfractions, as would be expected from the results of tracer studies. A role for the fatty acyl composition of high density lipoproteins in the determination of high density lipoprotein subfraction distribution is proposed.

Rate zonal ultracentrifugation of plasma samples from ten healthy age-matched volunteers (five males, five females) indicated that the high density lipoprotein subfraction ratio (HDL2:HDL3) in females was significantly higher than in males. The cause of this phenomenon was investigated by simultaneous examination of the metabolism of the major HDL apoproteins (apoA-I and apoA-II) in both groups. The results show that there is no significant sex-related difference in the plasma pool size, fractional catabolic rate, or synthetic rate of either apoprotein. We conclude that the increased HDL2:HDL3 ratio in females versus males does not derive from measurable differences in the metabolic handling of either apoprotein.

The effect of a rat high-density lipoprotein subfraction (HDL2) on the synthesis of bile salts by rat hepatocyte monolayers prepared from rats fed a diet containing cholestyramine, was investigated. The synthesis of bile salts as measured by radioimmunoassay of conjugated cholic, chenodeoxycholic and beta-muricholic acids was significantly increased when hepatocytes were incubated with a physiological concentration (500 micrograms HDL2 protein X ml-1) of HDL2.