Protein Biomarkers and Neuroproteomics Characterization of Microvesicles/Exosomes from Human Cerebrospinal Fluid Following Traumatic Brain Injury

Abstract

Recently, there have been emerging interests in the area of microvesicles and exosome (MV/E) released from brain cells in relation to neurodegenerative diseases. However, only limited studies focused on MV/E released post-traumatic brain injury (TBI) as they highlight on the mechanistic roles of released proteins. This study sought to examine if CSF samples from severe TBI patients contain MV/E with unique protein contents. First, nanoparticle tracking analysis determined MV/E from TBI have a mode of 74-98 nm in diameter, while control CSF MV/E have a mode of 99-104 nm. Also, there are more MV/E were isolated from TBI CSF (27.8-33.6 × 10/mL) than from control CSF (13.1-18.5 × 10/mL). Transmission electron microscopy (TEM) visualization also confirmed characteristic MV/E morphology. Using targeted immunoblotting approach, we observed the presence of several known TBI biomarkers such as αII-spectrin breakdown products (BDPs), GFAP, and its BDPs and UCH-L1 in higher concentrations in MV/E from TBI CSF than their counterparts from control CSF. Furthermore, we found presynaptic terminal protein synaptophysin and known exosome marker Alix enriched in MV/E from human TBI CSF. In parallel, we conducted nRPLC-tandem mass spectrometry-based proteomic analysis of two control and two TBI CSF samples. Ninety-one proteins were identified with high confidence in MV/E from control CSF, whereas 466 proteins were identified in the counterpart from TBI CSF. MV/E isolated from human CSF contain cytoskeletal proteins, neurite-outgrowth related proteins, and synaptic proteins, extracellular matrix proteins, and complement protein C1q subcomponent subunit B. Taken together, following severe TBI, the injured human brain released increased number of extracellular microvesicles/exosomes (MV/E) into CSF. These TBI MV/E contain several known TBI biomarkers and previously undescribed brain protein markers. It is also possible that such TBI-specific MV/E might contain cell to cell communication factors related to both cell death signaling a well as neurodegeneration pathways.

Current and Future Applications of Biomedical Engineering for Proteomic Profiling: Predictive Biomarkers in Neuro-Traumatology

“…The 16 articles selected fall into two main categories: 8 clinical papers and 8 laboratory studies on animal models of TBI [ 11 , 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 , 25 , 26 ]. Briefly, the 8 clinical studies identified have described four main technologies for identification of predictive biomarkers: (a) Microvescicle/Exosome, (b) MicroRNA, (c) Mass Spectroscopy, and (d) Multiplexing and Immunoassays [ 11 , 12 , 13 , 14 , 15 , 16 , 17 , 18 ].…”

Proteomic changes in traumatic brain injury: experimental approaches

“…This study examined exosomes, isolated from human CSF following TBI, using a proteomics approach [ 8 ▪▪ ]. Exosomes or microvesicles/exosomes (MV/E) are released from brain cells during disease and injury.…”

Complement C5a Induces Pro-inflammatory Microvesicle Shedding in Severely Injured Patients

“…These packages actively contribute in inflammatory processes by interacting with immune cells, epithelial and endothelial cells, and bridging innate and adaptive immunity ( 19 , 20 ). Several studies showed that MV numbers and their origin are altered in trauma and sepsis ( 21 – 23 ). Patients with major burn injury showed elevated numbers of granulocyte- and monocyte- derived MVs, and this increase was associated with injury severity ( 24 ).…”