Endocytosis caveolae-mediated

LDH-FITC is well overlapped with red fluorescent clathrin-TR, but not with caveolin-1-TR (Figure 13.10). This is dear evidence that clathrin-mediated endocytosis is the prindpal mechanism for the cellular internalization of LDH particles. Caveolae-mediated endocytosis, if any, seems not to be responsible for LDH uptake. 

Rejman J, Oberle V, Zuhom IS, Hoekstra D (2004) Size-dependent internalization of particles via the pathways of clathrin- and caveolae-mediated endocytosis. Biochem J 377 159-169... 

Rejman J, Bragonzi A, Conese M (2005) Role of clathrin- and caveolae-mediated endocytosis in gene transfer mediated by lipo- and polyplexes. Mol Ther 12 468 174... 

Fig. 2.2 Cellular internalisation pathways proposed for carbon nanotubes (CNTs) (A) phagocytosis (B) membrane piercing by passive diffusion (C) caveolae-mediated endocytosis and (D) clathrin-mediated endocytosis... 

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). 

In addition to the well characterized roles of clathrin-caveolae-mediated endocytosis and macropinocytosis/phagocytosis, an ill-defined route of nonclathrin-noncaveolae mediated endocytosis still exists (31,32). It seems that all of the until now poorly understood mechanisms of internalization can be summarized in this topic. 

Caveolae-Mediated Endocytosis and Caveolae-Like Endocytosis Pharmacological Inhibitors... 

In many studies, folate or folic acid (5nM) is applied to study caveolae-mediated endocytosis resp. potocytosis (27,119,120). 

Different incubation times and concentrations of applied marker might lead to different patterns of distribution. For example, LysoTracker Red is only selective for lysosomes when applied in low concentrations for a short time (10 minutes prior to imaging). The same has been described for other markers such as EGF and Tfn (as described in the section Caveolae-Mediated Endocytosis ). 

Keep in mind that exocytosis might occur during sample preparation (e.g., preparing cells for flow cytometric analysis). After the experiment (and prior to analysis) cells should be kept below 4°C to block all active processes (such as exocytosis). Exocytosis might also be blocked with NEM (see section Caveolae-Mediated Endocytosis and Caveolae-Like Endocytosis Pharmacological Inhibitors ). 

Gene delivery systems can distribute plasmids to the desired target cells, after which the plasmid is internalized into the cell by a number of mechanisms, such as adsorptive endocytosis, receptor-mediated endocytosis, micropinocytosis, caveolae-mediated endocytosis and phagocytosis (see Section 1.3.3.2). The intracellular fate of plasmids depends on the means by which they are internalized and translocated to the cytoplasms and then to the nucleus. In coated-pit endocytosis, DNA complexes first bind to the cell surface, then migrate to clathrin-coated pits about 150 ran in diameter and are internalized from the plasma membrane to form coated vesicles. 

Cell-surface receptors are involved in both phagocytosis and pinocytosis. At least four distinct mechanisms of pinocytosis have been characterized macropinocytosis, clathrin-mediated endocytosis, raft/caveolae-mediated endocytosis, and clathrin-and caveolae-independent endocytosis (1). Selected receptor-mediated aspects of these mechanisms are outlined below. 

Several mechanisms of endocytotic pathways exist. These include clathrin-mediated endocytosis, caveolae-mediated endocytosis, macropinocytosis, and non-clathrin receptor-mediated endocytosis (Table 4). All of these pathways can be exploited for drug trafficking into resistant cancer cells. For example, the... 

Caveolae-mediated endocytosis is involved in viral transfection. This route can therefore be used for the delivery of oncolytic genetic materials by a viral vector [120]. Macropinocytosis is a relatively non-specific process which allows uptake of large particles up to the micron size range [121]. It is likely useful for the delivery of systems like solid lipid nanoparticles and PLN, which are... 

Fig. 1 Proposed model for the internalization of PTD-conjugated macromolecules into cells. Interaction of positively charged PTDs with negatively charged proteoglycans and glycosa-minoglycans plays an important role in the internalization. The electrostatic interaction is followed by energy- and temperature-dependent endocytotic pathways. This involves phago-cytotic and pinocytotic pathways clathrin-mediated endocytosis, caveolae-mediated endocytosis, and macropinocytosis...

A typical feature of caveola-mediated endocytosis is the formation of non-coated invaginations composed of detergent-resistant membrane components rich in cholesterol and sphingolipids, known as lipid rafts [56]. The importance of the caveola-mediated mechanism in the PTD-mediated internalization was confirmed in an experiment where the cellular uptake of Tat peptide was affected by drugs that either disrupt lipid rafts or alter caveolar trafficking [57]. Moreover, Tat-PTD-fused protein showed colocalization with a marker of caveolar uptake, caveolin, further strengthening the importance of the mechanism in the PTD-mediated internalization [57]. 

Eukaryotic cells take up extracellular material by a variety of different mechanisms that are collectively termed as endocytosis. Endocytosis is an uptake mechanism for molecules from the extracellular area by invagination of the cell membrane.There are four routes for the uptake of nanoparticles clathrin-mediated endocytosis, caveolae-mediated endocytosis, macropinocytosis and clathrin- and caveolae-independent endocytosis. 

Besides clathrin-mediated cell uptake, caveolae-mediated endocytosis is another well-investigated uptake mechanism. The caveolae pathway is particularly attractive for the delivery of proteins and nucleic acids because these vesicles have neutral pH and the lysosomes, where degradation takes place, are bypassed [8,13]. Nanoparticles... 

Rejman J, Conese M, Hoekstra D (2006) Gene transfer by means of lipo- and polyplexes role of clathrin and caveolae-mediated endocytosis. J Liposome Res 16 237-247. doi 10.1080/ 08982100600848819... 

Cell culture substrate can also the influence gene delivery by affecting the occurrence of endocytosis and, thereby, the uptake of DNA-bound NPs by the cells. Hsu et al. [139] observed that the culture of MSCs on chitosan or HA-modified chitosan membranes increased the intracellular uptake of iron oxide NPs ( 5 nm) as well as naked DNA (3.3 kb, 5 mn) by more than fivefold. The increased internalization of NPs was associated with an increase in clathrin-mediated endocytosis on chitosan ( 50%) and in caveolae-mediated endocytosis on chitosan-HA ( 30-40%). In the case of naked DNA, but not iron oxide NPs, macropinocytosis also occurred on both substrates. 

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