Kruth Howard S, Jones Nancy L, Huang Wei, Zhao Bin, Ishii Itsuko, Chang Janet, Combs Christian A, Malide Daniela, Zhang Wei-Yang
Section of Experimental Atherosclerosis, NHLBI, National Institutes of Health, Bethesda, Maryland 20892-1422, USA.
J Biol Chem. 2005 Jan 21;280(3):2352-60. doi: 10.1074/jbc.M407167200. Epub 2004 Nov 8.
Previously, we reported that fluid-phase endocytosis of native LDL by PMA-activated human monocytederived macrophages converted these macrophages into cholesterol-enriched foam cells (Kruth, H. S., Huang, W., Ishii, I., and Zhang, W. Y. (2002) J. Biol. Chem. 277, 34573-34580). Uptake of fluid by cells can occur either by micropinocytosis within vesicles (<0.1 microm diameter) or by macropinocytosis within vacuoles ( approximately 0.5-5.0 microm) named macropinosomes. The current investigation has identified macropinocytosis as the pathway for fluid-phase LDL endocytosis and determined signaling and cytoskeletal components involved in this LDL endocytosis. The phosphatidylinositol 3-kinase inhibitor, LY294002, which inhibits macropinocytosis but does not inhibit micropinocytosis, completely blocked PMA-activated macrophage uptake of fluid and LDL. Also, nystatin and filipin, inhibitors of micropinocytosis from lipid-raft plasma membrane domains, both failed to inhibit PMA-stimulated macrophage cholesterol accumulation. Time-lapse video phase-contrast microscopy and time-lapse digital confocal-fluorescence microscopy with fluorescent DiI-LDL showed that PMA-activated macrophages took up LDL in the fluid phase by macropinocytosis. Macropinocytosis of LDL depended on Rho GTPase signaling, actin, and microtubules. Bafilomycin A1, the vacuolar H+-ATPase inhibitor, inhibited degradation of LDL and caused accumulation of undegraded LDL within macropinosomes and multivesicular body endosomes. LDL in multivesicular body endosomes was concentrated >40-fold over its concentration in the culture medium consistent with macropinosome shrinkage by maturation into multivesicular body endosomes. Macropinocytosis of LDL taken up in the fluid phase without receptor-mediated binding of LDL is a novel endocytic pathway that generates macrophage foam cells. Macropinocytosis in macrophages and possibly other vascular cells is a new pathway to target for modulating foam cell formation in atherosclerosis.
此前，我们报道过，佛波酯（PMA）激活的人单核细胞衍生巨噬细胞对天然低密度脂蛋白（LDL）进行液相内吞作用，可将这些巨噬细胞转化为富含胆固醇的泡沫细胞（Kruth, H. S., Huang, W., Ishii, I., and Zhang, W. Y. (2002) J. Biol. Chem. 277, 34573 - 34580）。细胞摄取液体可通过小泡（直径<0.1微米）内的微胞饮作用，或通过液泡（直径约0.5 - 5.0微米）内的巨胞饮作用，这些液泡称为巨吞饮小体。当前研究已确定巨胞饮作用是液相LDL内吞作用的途径，并确定了参与此LDL内吞作用的信号传导和细胞骨架成分。磷脂酰肌醇3激酶抑制剂LY294002可抑制巨胞饮作用，但不抑制微胞饮作用，它完全阻断了PMA激活的巨噬细胞对液体和LDL的摄取。此外，制霉菌素和菲律宾菌素是脂质筏质膜结构域微胞饮作用的抑制剂，二者均未能抑制PMA刺激的巨噬细胞胆固醇积累。延时视频相差显微镜和使用荧光DiI-LDL的延时数字共聚焦荧光显微镜显示，PMA激活的巨噬细胞通过巨胞饮作用摄取液相LDL。LDL的巨胞饮作用依赖于Rho GTPase信号传导、肌动蛋白和微管。液泡H⁺-ATP酶抑制剂巴弗洛霉素A1抑制LDL的降解，并导致未降解的LDL在巨吞饮小体和多泡体内涵体内积累。多泡体内涵体中的LDL浓度比其在培养基中的浓度浓缩了40多倍，这与巨吞饮小体通过成熟为多泡体内涵体而收缩一致。液相摄取且无受体介导的LDL结合情况下的LDL巨胞饮作用是一种产生巨噬细胞泡沫细胞的新型内吞途径。巨噬细胞以及可能其他血管细胞中的巨胞饮作用是调节动脉粥样硬化中泡沫细胞形成的一个新的靶向途径。
