Label-free quantification proteomics reveals novel calcium binding proteins in matrix vesicles isolated from mineralizing Saos-2 cells

doi:10.5582/bst.2013.v7.3.144

Biosci Trends

2013

DOI: 10.5582/bst.2013.v7.3.144

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Label-free quantification proteomics reveals novel calcium binding proteins in matrix vesicles isolated from mineralizing Saos-2 cells

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Cited by 4 publications

References 37 publications

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Exosome enrichment of human serum using multiple cycles of centrifugation

2015

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In this work, we compared the use of repeated cycles of centrifugation at conventional speeds for enrichment of exosomes from human serum compared to the use of ultracentrifugation. After removal of cells and cell debris, a speed of 110,000×g or 40,000×g was used for the ultracentrifugation or centrifugation enrichment process, respectively. The enriched exosomes were analyzed using the BCA assay, 1-D gel separation, transmission electron microscopy, Western blotting, and high resolution LC-MS/MS analysis. It was found that a five cycle repetition of ultracentrifugation or centrifugation is necessary for successful removal of non-exosomal proteins in the enrichment of exosomes from human serum. More significantly, 5×centrifugation enrichment was found to provide similar or better performance than 5×ultracentrifugation enrichment in terms of enriched exosome protein amount, Western blot band intensity for detection of CD-63 and numbers of identified exosome-related proteins and CD proteins. A total of 478 proteins were identified in the LC-MS/MS analyses of exosome proteins obtained from 5×ultracentrifugations and 5×centrifugations including many important CD membrane proteins. The presence of previously reported exosome-related proteins including key exosome protein markers demonstrates the utility of this method for analysis of proteins in human serum.

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Exosome enrichment of human serum using multiple cycles of centrifugation

2015

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Matrix vesicles: Are they anchored exosomes?

Shapiro

Landis

Risbud

2015

Bone

153

131

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Numerous studies have documented that matrix vesicles are unique extracellular membrane-bound microparticles that serve as initial sites for mineral formation in the growth plate and most other vertebrate mineralizing tissues. Microparticle generation is not confined to hard tissues, as cells in soft tissues generate similar structures; numerous studies have shown that a common type of extracellular particle, termed an exosome, a product of the endosomal pathway, shares many characteristics of matrix vesicles. Indeed, analyses of size, morphology and lipid and protein content indicate that matrix vesicles and exosomes are homologous structures. Such a possibility impacts our understanding of the biogenesis, processing and function of matrix vesicles (exosomes) in vertebrate hard tissues and explains in part how cells control the earliest stages of mineral deposition. Moreover, since exosomes influence a spectrum of functions, including cell-cell communication, it is suggested that this type of microparticle may provide a mechanism for the transfer of signaling molecules between cells within the growth plate and thereby regulate endochondral bone development and formation.

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Core level regulatory network of osteoblast as molecular mechanism for osteoporosis and treatment

Yuan

Zhu³

et al. 2016

Oncotarget

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To develop and evaluate the long-term prophylactic treatment for chronic diseases such as osteoporosis requires a clear view of mechanism at the molecular and systems level. While molecular signaling pathway studies for osteoporosis are extensive, a unifying mechanism is missing. In this work, we provide experimental and systems-biology evidences that a tightly connected top-level regulatory network may exist, which governs the normal and osteoporotic phenotypes of osteoblast. Specifically, we constructed a hub-like interaction network from well-documented cross-talks among estrogens, glucocorticoids, retinoic acids, peroxisome proliferatoractivated receptor, vitamin D receptor and calcium-signaling pathways. The network was verified with transmission electron microscopy and gene expression profiling for bone tissues of ovariectomized (OVX) rats before and after strontium gluconate (GluSr) treatment. Based on both the network structure and the experimental data, the dynamical modeling predicts calcium and glucocorticoids signaling pathways as targets for GluSr treatment. Modeling results further reveal that in the context of missing estrogen signaling, the GluSr treated state may be an outcome that is closest to the healthy state.

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Label-free quantification proteomics reveals novel calcium binding proteins in matrix vesicles isolated from mineralizing Saos-2 cells

Cited by 4 publications

References 37 publications

Exosome enrichment of human serum using multiple cycles of centrifugation

Exosome enrichment of human serum using multiple cycles of centrifugation

Matrix vesicles: Are they anchored exosomes?

Core level regulatory network of osteoblast as molecular mechanism for osteoporosis and treatment

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