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Figure 1. Electron micrographs of exosomes released from the human colon carcinoma cell line LIM1215. Exosomes were isolated from culture media by differential centrifugation as described in [139]. (A) Uranyl acetate staining. (B) Anti-A33 immunogold/uranyl acetate staining. Typical exosome yield is approximately 50 µg/5 × 108 cells/24 h.
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Figure 2. Exosome generation. A receptor is internalized following ligand binding and is trafficked to early endosomes. In early endosomes, proteins are either recycled to the plasma membrane or sequested in ILVs of the MVBs. ILVs of MVBs are generated by budding from the limiting membrane into the lumen of endosomes [111]. MVBs can be either degradative (evolving into lysosomes) or it can be exocytic (fuse with plasma membrane to release its contents). The degradative MVBs can be generated by the endosomal sorting complex required for transport or ubiquitination [5,112,113]. The exocytic MVBs fuse with the plasma membrane to release exosomes to the microenvironment. Exosomes display the same surface topology as the plasma membrane with extracellular domains of proteins at the surface, and enclosing cytoplasmic contents in their lumen. According to a model proposed by Greco and colleagues [114] , released exosomes can be internalized by neighboring cells and act as a potential vehicle for spreading morphogens through epithelia; alternatively, membrane protruding from secreting cells may be phagocytosed by a neighboring cell and fragmentation of this internalized membrane may lead to the formation of MVBs in neighboring cells. ILV: Internal luminal vesicle; MVB: Multivesicular body. Adapted from [115].
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Figure 3. Overlap of three exosome studies that have identified micro (mi)RNAs. The numbers outside each circle represent the total number of miRNAs identified in the respective study. Interestingly, only eight miRNAs are found to be in common among the three exosome studies.
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Figure 4. Recent instrument developments in mass spectrometry has led to the utilization of the dual LTQ-Orbitrap mass spectrometer for high sensitivity and high mass-accuracy peptide sequencing using dual fragmentation capabilities of CID and ETD [116-120]. The LTQ-Orbitrap is superior in the detection and identification of peptides in a routine pipeline for the analysis of proteotypic peptide derived from tryptic digests of proteins recovered from the protein complement of...