Characterization And Isolation Of A Novel Type Of Cell Surface Microvesicles

Cell surface microvesicles are cell-bond spherical projection,They will be presented on the cell surface of the membrane when adherent cells spread and their diameter is 1um.Currently,some reports of the vesicle-like structure on membrane surface is that cell surface microvesicles are precursors of endothelial microparticles,but it is just a kind of inference without direct evidences;At the same time,there are no efficient methods to research this structure deeply,because of no effective methods to separate and purify them.So it is necessary to distinguish and clarify that whether cell surface microvesicles are precursors of endothelial microparticles or independent structures and physiological functions.Simultaneously,it is important to establish an effective method to separate and purify cell surface microvesicles in order to lay the foundation of the structure and function of fellow-up study.Using HUVEC and HepG-2 as the analysis cell models to research cell surface microvesicles in this study.Firstly,our results determine that cell surface microvesicles are cell-bonded and cell surface membrane structures by confocal microscope and scanning electron microscope;The results of immunostaining of specific markers indicate that these projections are not the results of mitochondria and multivesicular bodies due to they set up the membranes.These microvesicles do not belong to the extracellular microparticles,exosomes and apoptotic bodies through the results of immunostaining of specific markers,such as CD31,CD146,CD63 and PS.Experiments also found that cell surface microvesicles can be dissolved by ?70% ethanol and chloroform and methanol mixture and can be protected by osmium tetroxide under such conditions.These indicate that cell surface microvesicles are the structures contained lipid compositions.Furthermore,these surface microvesicles may contain FAK, GM1, GM3 or caveolin-1,but there are not specific.Further strong evidences show that cell surface microvesicles have strong resistance to Triton X-100, SDS and saponin,this is a unique special character of these microvesicles.Most importantly, using their unique physical and chemical properties,we establish a preliminary method to isolate cell surface microvesicles,which offers the potential for the molecular characteristics of membrane structure and composition analysis.This paper provides a solid and reliable evidence that cell surface microvesicles are neither bulbous protrusions caused by intracellular organelles such as mitochondria and multivesicular bodies nor precursors of extracellular vesicles such as extracellular microparticles,exosomes and apoptotic bodies, they are cell-bond spherical microvesicles and have the unique lipid compositions and physical properties.And more importantly,we establish a simple and effective method to separate and purify these vesicles in this paper,this makes its follow-up study of composition and structure possible.The illumination of these vesicles will create a new field on cell biology,and the establish of separation method will promote its research.Thus, this research is extremely important.