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Updated: Jul 16, 2026
Isolation and Analysis of Plasma Lipoproteins by Ultracentrifugation
Published on: January 28, 2021
Apolipoprotein A-I complexed with phospholipid promotes hepatic lipoprotein and apolipoprotein secretion in the
1Department of Medicine, University of Louisville, School of Medicine, KY 40292, USA.
Background:
Apolipoprotein A-I within high density lipoprotein (HDL) plays a significant role in the process of reverse cholesterol transport from peripheral tissues to the liver. However, additional roles are not well defined for it in hepatic cholesterol metabolism. We have previously shown in the hamster that dietary cholesterol supplementation resulted in enhancement of apolipoprotein A-I (Apo A-I) in secreted nascent hepatic very low density lipoprotein (VLDL), suggesting that apolipoprotein A-I itself may play a role in hepatic lipoprotein secretion.
Methods:
Using the isolated hamster liver with Apolipoprotein A-I perfusion, we then examined the hypothesis that Apo A-I alone or in association with phosphotidylcholine (PC) i.e., Apo A-I/PC as a HDL-like particle, has effects upon hepatic lipoprotein and bile secretion. Ultracentrifugation was performed on perfusate samples at 3 hours on control vs treated livers (Apo A-I/PC, Apo A-I, or PC) to access lipid and protein concentration in VLDL, low density lipoprotein (LDL) and HDL. Four to thirty percent gradient SDS polyacrylamide electrophoresis (PAGE) and Western blot analysis were used on delipidated lipoprotein fractions and microsomes to assess apolipoproteins Apo B, A-I, II, and E.
Results:
We found that perfusion of reconstituted HDL vesicles containing human apolipoprotein A-I and PC (Apo A-I/PC) 10 mg and 10 mg, respectively, in 22 mL for 3 hours into isolated hamster liver increased cholesterol (CH) and triglyceride (TG) components in secreted HDL; 45- and 6-fold, and in LDL; 15- and 2-fold, respectively. No significant changes occurred in VLDL or in biliary lipids. Concomitantly, Apo A-I/PC perfusion increased Apo E and Apo A-II and HDL and Apo B in LDL, while Apo E decreased in VLDL. Apo A-I/PC perfusion did not change the apolipoprotein content of hepatic microsomes of the perfused liver. Perfusion of apolipoprotein A-I (without PC) or PC (without apolipoprotein A-I) had none of these effects.
Conclusion:
These results indicate that the perfused discoidal apolipoprotein A-I/PC particle affects hepatic lipoprotein assembly and secretion, whereby both lipid and apolipoprotein components are enhanced in secreted HDL and LDL of hepatic origin.
