Liposomes endocytosis

The involvement of the actin cytoskeleton in liposome endocytosis is studied by using cytochalasins, latrunculin, or toxin C2 to polymerize actin filaments. For a review on actin assembly and endocytosis, see Ref. (135). 

They encapsulated poly(MA-CDA) into mannan-coated liposomes and evaluated superoxide production from mouse macrophages. The activity was three- to five-fold high compared with uncapsulated poly(MA-CDA) itself [5,11], suggesting that an increased incorporation of the polymer by the receptor-mediated endocytosis mediated the higher biological activity.. 

Nakae, D., Yoshiji, H., Amanuma, T., Kinugasa, T., Farber, J.L. andKonishi, Y. (1990). Endocytosis-independent uptake of liposome-encapsulated superoxide dismutase prevents the killing of cultured hepatocytes by tert-butyl hydroperoxide. Arch. Biochem. Biophys. 279, 315-319. 

Endocytosis of the vesicle by the cell. The entire liposomal contents are made available to the cell. 

Matthay, K.K., Heath, T.D., Badger, C.C., Bernstein, I.D., and Papahadjopoulos, D. (1986) Antibody-directed liposomes Comparison of various ligands for association, endocytosis and drug delivery. Cancer Res. 46, 4904. 

Lee, K.-D., Hong, K. and Papahadjopoulos, D. (1992). Recognition of liposomes by cells in vitro binding and endocytosis mediated by specific lipid headgroups and surface charge density, Biochim. Biophys. Acta, 1103, 185-197. 

Lee, K.-D., Nir, S. and Papahadjopoulos, D. (1993). Quantitative analysis of liposome-cell interactions in vitro rate constants of binding and endocytosis with suspension and adherent J774 cells and human monocytes, Biochemistry, 32, 889-899. 

Miller CR, Bondurant B, McLean SD, et al. Liposome-cell interactions in vitro effect of liposome surface charge on the binding and endocytosis of conventional and sterically stabilized liposomes. Biochemistry 1998 37 12875. 

Fretz MM, et al. OVCAR-3 cells internalize TAT-peptide modified liposomes by endocytosis. Biochim Biophys Acta 2004 1665 48. 

Wrobel I, Collins D. Fusion of cationic liposomes with mammalian cells occurs after endocytosis. Biochim Biophys Acta 1995 1235(2) 296-304. 

Integrin receptor-binding peptides have been used to enhance liposome binding, uptake, and expression (25,47 9). The inclusion of an 0(5pi integrin-targeted peptide into a liposomal complex enhanced transfection efficiency four- to five-fold in Jurkat cells and 10- to 13-fold in TF-1 cells (48). Confocal and electron microscopy revealed that the mechanism of cell entry conferred by RGD peptides on liposomes is predominantly by clathrin-coated endocytosis rather than by phagocytosis (50). 

Figure 2 Proposed pathways for liposomal entry into the cell enhanced by peptides. These include direct cell entry suggested as the mechanism of entry by cell-penetrating peptides and receptor-mediated endocytosis by caveolae- and clathrin-dependent endocytosis.

It has also been proposed that CPP are taken up by clathrin-independent mechanisms such as caveolin-mediated endocytosis (100). TAT-containing liposomes were colocalized with caveolin 1, a marker for caveolar endocytosis, but not with markers for clathrin (101). An inhibitor of caveolin and nystatin reduced TAT peptide reporter in HepG2 and CHO cells by 50% (80). However, it was pointed out by Brooks et al. that HepG2 cell lines do not contain caveolin 1 (102,103). The caveolin pathways relevance to CPP uptake may depend on the cell type because nystatin inhibits... 

Previous work has shown that the majority of cells internalize liposomes through an endocytic pathway (4,5). There are multiple pathways for internalization involving vesicles of 50 300 nm in diameter. These include clathrin-mediated endocytosis, caveolae-mediated endocytosis, phagocytosis, macropinocytosis, and nonclathrin- noncaveolae-dependent endocytosis (6). 

There are also multiple pathways for liposomes following cellular uptake. They may be delivered to lysosomes, recycled out of the cell, involved in transcytotic passage across an epithelial barrier, or delivered to other cellular compartments such as the Golgi network. Each route offers opportunities for selective delivery of macromolecular drugs and nanosized drugs so the need to comprehend endocytic pathways has never been more apparent (7). Figure 1 summarizes the different pathways of endocytosis. 

Figure 1 Endocytosis of liposomes five different routes into the cell. Multiple pathways can be used by the cell to internalize liposomes. Besides the well-characterized clathrin-mediated endocytosis, other pathways can be applied by the cell. Possible alternative pathways include phagocytosis or macropinocytosis—two pathways that internalize by an actin-driven protuberance of the plasma membrane. Other routes include the involvement of caveolae where substances are taken up into the cell bypass the traditional endosome/lysosome system (particles might escape from being degraded in lysosomes). Finally there exists an ill-defined mechanism that is neither mediated by caveolae nor by clathrin. In a single cell type, two or more of these mechanisms can coexist. Source Adapted from Ref 8.

Cholera Toxin Subunit B. a marker for caveolae-dependent endocytosis. partly co-localizes with liposomal FITC>dextran indicating that pH-sensitive liposomes are taken up-at least to a certain extend— via caveotae pathway... 

The different mechanisms of uptake and cellular processing were studied by the use of different inhibitors, which are summarized and reviewed in T able 1. Uptake mechanisms can be investigated by looking for colocalization of fluorescently labeled liposomes and labeled markers for endocytosis. This section describes how to study the initial mode of internalization, whereas... 

Incubation at 4°C (see section Energy Dependence on Liposome Uptake and Fusion with Cell Membranes ) and a block of metabolic activity (see section Metabolic Activity ) might also be used to block endocytosis and to detect cellular association or fusion. 

Lakkaraju A, Rahman YE, Dubinsky JM. Low-density lipoprotein receptor-related protein mediates the endocytosis of anionic liposomes in neurons. J Biol Chem 2002 277(17) 15085-15092. 

Yoshimura T, Shono M, Imai K, Hong K. Kinetic analysis of endocytosis and intracellular fate of liposomes in single macrophages. J Biochem (Tokyo) 1995 117(1) 34 41. 

Straubinger RM, Papahadjopoulos D, Hong KL. Endocytosis and intracellular fate of liposomes using pyranine as a probe. Biochemistry 1990 29(20) 4929-4939. 

Daleke DL, Hong K, Papahadjopoulos D. Endocytosis of liposomes by macrophages binding, acidification and leakage of liposomes monitored by a new fluorescence assay. Biochim Biophys Acta 1990 1024(2) 352-366. 

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