Abstract
The mechanism for the assembly of HDL with cellular lipid by ABCA1 and helical apolipoprotein was investigated in hepatocytes. Both HepG2 cells and mouse primary culture hepatocytes produced HDL with apolipoprotein A-I (apoA-I) whether endogenously synthesized or exogenously provided. Probucol, an ABCA1 inactivator, inhibited these reactions, as well as the reversible binding of apoA-I to HepG2. Primary cultured hepatocytes of ABCA1-deficient mice also lacked HDL production regardless of the presence of exogenous apoA-I. HepG2 cells secreted apoA-I into the medium even when ABCA1 was inactivated by probucol, but it was all in a free form as HDL production was inhibited. When a lipid-free apoA-I-specific monoclonal antibody, 725-1E2, was present in the culture medium, production of HDL was suppressed, whether with endogenous or exogenously added apoA-I, and the antibody did not influence HDL already produced by HepG2 cells. We conclude that the main mechanism for HDL assembly by endogenous apoA-I in HepG2 cells is an autocrine-like reaction in which apoA-I is secreted and then interacts with cellular ABCA1 to generate HDL.
Highlights
The mechanism for the assembly of HDL with cellular lipid by ABCA1 and helical apolipoprotein was investigated in hepatocytes
Cholesterol content in the HDL fraction (d Ͼ 1.063) was increased by 2-fold, and the VLDL-LDL fraction (d Ͻ 1.063) was insignificantly decreased. These results indicated that the system is functional both in HepG2 cells and in mouse hepatocytes for generation of the HDL-size lipoprotein by helical apolipoprotein
ABCA1 was resistant against degradation in the probucol-loaded HepG2 cells (Fig. 7C, D) in the same manner as we demonstrated in fibroblasts [11]
Summary
The mechanism for the assembly of HDL with cellular lipid by ABCA1 and helical apolipoprotein was investigated in hepatocytes. The main site for HDL production is generally thought to be the liver and intestine, where the cells synthesize helical apolipoprotein, mainly apolipoprotein A-I (apoA-I), and produce HDL, presumably upon the interaction of this apolipoprotein with its own ABCA1 by removing cellular lipid [6,7,8]. It is unclear in which step of the apoA-I production and secretion this reaction takes place for the assembly of HDL.
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