Proteomic analysis of urine exosomes by multidimensional protein identification technology (MudPIT)

Abstract

Exosomes are membrane vesicles that are secreted by cells upon fusion of multivesicular bodies with the plasma membrane. Exosomal proteomics has emerged as a powerful approach to understand the molecular composition of exosomes and has potential to accelerate biomarker discovery. Different proteomic analysis methods have been previously employed to establish several exosome protein databases. In this study, TFE solution phase digestion was compared with in-gel digestion and found to yield similar results. Proteomic analysis of urinary exosomes was performed by multidimensional protein identification technology (MudPIT) after TFE digestion. 3280 proteins were identified from nine human urine samples with 31% overlap among nine samples. Gene ontology (GO) analysis, coupled with detection of all of the members of ESCRT machinery complex, supports the multivesicular origin of these particles. These results significantly expand the existing database of urinary exosome proteins. Our results also indicate that more than 1000 proteins can be detected from exosomes prepared from as little as 25 mL of urine. This study provides the largest set of proteins present in human urinary exosome proteomes, provides a valuable reference for future studies, and provides methods that can be applied to exosomal proteomic analysis from other tissue sources.

Exosomes Derived from HIV-1-infected Cells Contain Trans-activation Response Element RNA

“…Exosomes contain lipids, proteins, and nucleic acids (mRNAs and miRNAs) 2 (5,6). The proteomic composition of exosomes has been well characterized (7)(8)(9)(10). Exosomes released into the intercellular space can fuse with multiple target cells and exert regulatory influences on the target cell (11)(12)(13)(14)(15).…”

Activation of the Endogenous Renin-Angiotensin-Aldosterone System or Aldosterone Administration Increases Urinary Exosomal Sodium Channel Excretion

“…5 In the present study, we tested the hypothesis that RAAS activation during a lowsodium (LS) diet and during exogenous aldosterone infusion alters the urinary exosome proteomic profile. We extended these findings using a more targeted, sensitive, and quantitative approach to investigate the profile of ENaC and NCC, which are known to play an essential role in renal sodium and potassium homeostasis.…”

Extracellular presence/release of galectins from HTR-8/SVneo extravillous trophoblast cells*

“…The findings presented here are novel for gal-3 and gal-8, since no literature data regarding their presence in EVs of trophoblast origin have been reported so far. However, both gal-3 and gal-8 were found in EVs from different cell types or body fluids (Ogawa et al, 2008;Wang et al, 2012;Webber et al, 2014;Hurwitz et al, 2016).…”