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Modified LDLs are internalized by macrophages in part via macropinocytosis
1Department of Pathology, Wake Forest University School of Medicine, Winston-Salem, North Carolina 27157-1092, USA. njones@wfubmc.edu
Abstract:
Macrophage foam cell formation in vitro requires uptake of modified low density lipoproteins (LDL) such as acetylated LDL (AcLDL) and moderately oxidized LDL (OxLDL). Macrophages incubated with AcLDL and OxLDL, but not LDL, showed increased membrane ruffling as seen with time-lapse phase contrast video light microscopy. Modified LDLs stimulated circular membrane ruffles between 2 and 10 min after incubation. These membrane ruffles were readsorbed into the plasma membrane between 5 and 15 min later. Phase-bright macropinosomes formed at the base of the stimulated membrane ruffles. The fluid-phase marker lucifer yellow labeled the modified LDL stimulated macropinosomes. Modified LDLs stimulate fluid-phase uptake by 1.5-fold to threefold as measured with 14C-sucrose uptake. Transmission electron microscopy showed that gold conjugated AcLDL and OxLDL bound preferentially to membrane ruffles and were present in macropinosomes (diameter >0.2 pm) underneath these membrane ruffles. AcLDL and OxLDL were also present in clathrin-coated pits and endosomes. These studies suggest that modified lipoproteins stimulate macropinocytosis. AcLDL and OxLDL are partially internalized by macropinocytosis and partially internalized via clathrin-coated pit endocytosis.
Insights
Modified lipoproteins like acetylated LDL (AcLDL) and oxidized LDL (OxLDL) stimulate macrophages to form membrane ruffles and macropinosomes, enhancing fluid-phase uptake. This suggests modified LDLs are internalized via macropinocytosis and clathrin-coated pit endocytosis.
Area of Science:
- Cell Biology
- Lipid Metabolism
- Atherosclerosis Research
Background:
- Macrophage foam cell formation is crucial in atherosclerosis.
- Modified low-density lipoproteins (LDL), including acetylated LDL (AcLDL) and oxidized LDL (OxLDL), are implicated in this process.
- The cellular mechanisms of modified LDL uptake are not fully elucidated.
Purpose of the Study:
- To investigate the cellular mechanisms by which macrophages internalize modified LDLs.
- To determine if modified LDLs stimulate specific endocytic pathways.
- To understand the role of membrane ruffling and macropinocytosis in modified LDL uptake.
Main Methods:
- Time-lapse phase contrast video light microscopy to observe membrane ruffling.
- Use of fluid-phase markers (lucifer yellow) and radiolabeled sucrose (14C-sucrose) to measure fluid-phase uptake.
- Transmission electron microscopy (TEM) with gold-conjugated modified LDLs to visualize uptake pathways.
- Incubation of macrophages with native LDL, AcLDL, and OxLDL.
Main Results:
- AcLDL and OxLDL, but not native LDL, induced rapid membrane ruffling in macrophages.
- Macropinosomes formed at the base of stimulated membrane ruffles and were labeled by fluid-phase markers.
- Fluid-phase uptake increased 1.5- to threefold with modified LDLs.
- TEM confirmed that modified LDLs bound to membrane ruffles and were internalized via macropinosomes, clathrin-coated pits, and endosomes.
Conclusions:
- Modified lipoproteins, including AcLDL and OxLDL, actively stimulate macropinocytosis in macrophages.
- Macrophages internalize modified LDLs through a combination of macropinocytosis and clathrin-coated pit endocytosis.
- These findings elucidate key cellular mechanisms involved in modified LDL uptake and foam cell formation.