The uptake and degradation of matrix-bound lipoproteins by macrophages require an intact actin Cytoskeleton, Rho family GTPases, and myosin ATPase activity.
A key cellular event in atherogenesis is the interaction of macrophages with lipoproteins in the subendothelium. In vivo, these lipoproteins are bound to matrix and often aggregated, yet most cell-culture experiments explore these events using soluble monomeric lipoproteins. We hypothesized that the internalization and degradation of matrix-retained and aggregated low density lipoprotein (LDL) by macrophages may involve the actin-myosin cytoskeleton in a manner that distinguishes this process from the endocytosis of soluble LDL. To explore these ideas, we plated macrophages on sphingomyelinase-aggregated LDL bound to smooth muscle cell-derived matrix in the presence of lipoprotein lipase. The macrophages internalized and degraded the LDL, which was mediated partially by the LDL receptor-related protein. Cytochalasin D and latrunculin A, which block actin polymerization, markedly inhibited the uptake and degradation of matrix-retained LDL but not soluble LDL. Inhibition of Rho family GTPases by Clostridium difficile toxin B blocked the degradation of matrix-retained and aggregated LDL by >90% without any inhibition of soluble LDL degradation. However, specific inhibition of Rho had no effect, suggesting the importance of Rac1 and Cdc42. Degradation of matrix-retained, but not soluble, LDL was also blocked by inhibitors of tyrosine kinase, phosphatidylinositol 3-kinase, and myosin ATPase. These findings define fundamental cytoskeletal pathways that may be involved in macrophage foam cell formation in vivo but have been missed by the use of previous cell culture models.
Sakr
S W
SW
Department of Medicine, Columbia University, New York, New York 10032, USA.
Eddy
R J
RJ
Barth
H
H
Wang
F
F
Greenberg
S
S
Maxfield
F R
FR
Tabas
I
I
D000199:Actins / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism*; D008077:Lipoproteins, LDL / Q000378:metabolism*; D026503:Low Density Lipoprotein Receptor-Related Protein-1 / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D009218:Myosins / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D020741:rho GTP-Binding Proteins / Q000378:metabolism*
D000199:Actins; D008077:Lipoproteins, LDL; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases; D009218:Myosins; D020741:rho GTP-Binding Proteins