MACROPHAGE ACTIVATION AND LIPOPROTEIN METABOLISM

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We have demonstrated that the uptake of LDL-containing immune complexes (LDL-IC) by human monocyte-derived macrophages induces the transformation of these cells into foam cells and promotes their activation leading to a paradoxical increase in LDL receptor cell surface expression and to the release of cytokines. In this proposal we plan to analyze the molecular mechanisms responsible for the increase in LDL receptor cell surface expression. We will determine whether the increase in LDL receptor expression, observed in human macrophages stimulated by LDL-containing immune complexes (LDL-IC), is due to increased transcription of the LDL receptor gene, increased mRNA stability, increased mRNA translation, decreased LDL receptor protein degradation or increased translocation of the LDL receptor protein to the cell membrane. We will also investigate whether this increase in LDL receptor expression is secondary to a decrease in the intracellular cholesterol regulatory pool and if so, whether the depletion in this regulatory pool is secondary to enhanced sterol mobilization to the ACAT substrate pool. Furthermore, we will determine whether factors released during macrophage activation mediate or facilitate the increase in LDL receptor cell surface expression. We will also investigate whether the stimulation of LDL receptor cell surface expression is a consequence of the uptake of the LDL-containing immune complexes by a specific Fc receptor subtype (Fc-gamma-RI, Fc-gamma-RII or Fc-gamma-RIII). Since HMGCoA reductase activity and LDL receptor activity usually have a coordinate regulation, we will determine whether HMGCoA reductase activity is also increased in LDL-IC-stimulated macrophages. This project will involve cell culture, cell isolation and cell fractionation procedures, studies of intracellular lipid metabolism, receptor binding studies, immunoblotting and molecular biology techniques. Our overall objective is to provide insight into the mechanisms by which anti-LDL antibodies and macrophage activation may contribute to the development(acceleration of the arteriosclerotic process.