ApoA-IV is secreted on discrete HDL particles by the rat hepatoma cell line McA-RH7777 transfected with ApoA-IV cDNA

Abstract

In the present study, the synthesis and secretion of transfected apolipoprotein (apo) A-IV was investigated in rat hepatoma McA-RH7777, a cell line that does not express apoA-IV mRNA or protein. An expression plasmid that contained the rat apoA-IV cDNA was transfected into the cells; five stable transformants were selected that harbor different copy numbers of the apoA-IV construct and secrete different amounts of apoA-IV. Gel filtration column chromatography and density gradient ultracentrifugation, combined with gel electrophoresis and electron microscopy techniques, demonstrated that (1) the secreted apoA-IV associated mainly with high-density lipoproteins (HDLs) and only a trace amount of apoA-IV was associated with very-low-density lipoproteins; (2) overexpression of apoA-IV resulted in an increased number of disk-shaped structures (thickness, ≈8.0 nm and diameter, ≈22 nm); and (3) the electrophoretic mobilities of the apoA-IV-containing particles differed from those of apoA-I-containing HDL. Expression of apoA-IV exerted no discernible effect on the density distribution or the secretion efficiency of apoB-100. Additionally, secretion of apoB-100 and apoA-IV exhibited opposite responses to serum: apoB-100 secretion was stimulated eightfold after addition of serum, whereas apoA-IV secretion was inhibited by 40%. These results suggest that synthesis of apoA-IV may lead to the formation of a subclass of HDL with a different metabolic fate than that of lipoproteins containing either apoA-I or apoB.