Exosomal Composition, Biogenesis and Profiling Using Point-of-Care Diagnostics-Implications for Cardiovascular Disease

Abstract

Arteriosclerosis is an important age-dependent disease that encompasses atherosclerosis, in-stent restenosis (ISR), pulmonary hypertension, autologous bypass grafting and transplant arteriosclerosis. Endothelial dysfunction and the proliferation of vascular smooth muscle cell (vSMC)-like cells is a critical event in the pathology of arteriosclerotic disease leading to intimal-medial thickening (IMT), lipid retention and vessel remodelling. An important aspect in guiding clinical decision-making is the detection of biomarkers of subclinical arteriosclerosis and early cardiovascular risk. Crucially, relevant biomarkers need to be good indicators of injury which change in their circulating concentrations or structure, signalling functional disturbances. Extracellular vesicles (EVs) are nanosized membraneous vesicles secreted by cells that contain numerous bioactive molecules and act as a means of intercellular communication between different cell populations to maintain tissue homeostasis, gene regulation in recipient cells and the adaptive response to stress. This review will focus on the emerging field of EV research in cardiovascular disease (CVD) and discuss how key EV signatures in liquid biopsies may act as early pathological indicators of adaptive lesion formation and arteriosclerotic disease progression. EV profiling has the potential to provide important clinical information to complement current cardiovascular diagnostic platforms that indicate or predict myocardial injury. Finally, the development of fitting devices to enable rapid and/or high-throughput exosomal analysis that require adapted processing procedures will be evaluated.

Keywords: atheroscelorsis; endothelial (dys)function; exosome (vesicle); point of care diagnosis; stem cell repair mechanisms.

FIGURE 1

Exosome Biogenesis: Exosomes biogenesis is a complex process and can be conducted in an ESCRT-dependent and -independent manner. The internalisation of the plasma membrane initiates the formation of exosomes by endocytosis. The fusion of early endosomes results in the production of multivesicular bodies. Upon maturation, MVBs are directed to the plasma membrane via a tightly Rab GTPase dependent multistep process where they undergo a SNARE-mediated plasma membrane fusion event followed by secretion into the extracellular space.

FIGURE 2

Molecular components of vascular exosome: Exosomes are composed of a plasma membrane-derived phospholipid bilayer containing cytosol components from the cell of origin. The composition of the exosomes is dependent on the cell type of origin, the state of health of the source cell, and extracellular stimuli. There are various proteins, lipids, and miRNAs that are common to the majority of exosomes.

FIGURE 3

Exosome uptake by neighbouring cells: Schematic representation of exosome uptake by neighbouring cells. Once secreted into the extracellular space, exosomes can be taken up by neighbouring cells via two main processes i) exosomes are engulphed into the target cell via endocytosis. Once endocytosed exosomes fuse with the endocytic compartment delivering various cargo ii) exosomes dock onto the plasma membrane of the target cell and directly deliver their cargo into the cytoplasm of the target cell.

FIGURE 4

Endothelial exosomes in vascular health and disease: Under physiological conditions, endothelial cells release exosomes at low concentrations which facilitate the protection and vascular homeostasis. When exposed to pathological vascular stimuli, the content of these exosomes changes, resulting in a loss of vascular homeostasis, apoptosis, monocyte infiltration, and subsequent facilitation of the development of CVD.

FIGURE 5

Exosome-derived biomarkers for the diagnosis of CVD. Exosomes contain key potential biomarkers including various surface proteins, miRNAs and nucleic acids. Following isolation, exosomes can be measured, or further processed using exosome lysis before biomarker recognition to analyse contents.

FIGURE 6

Diagram of key steps/approaches for measuring exosome for the prediction of CVD. (A) Magnetic microbeads coated with specific antibodies enabled exosome isolation and addition of guanidine-based lysis buffer released the contents of enriched exosomes. Torque-actuated valves and permanent magnets were used to control the process. RNA is adsorbed on a glass-bead filter via electrostatic interactions and reagents are loaded to prepare for qPCR. (B) Images from scanning electron microscope of exosomes captured on antibody-coated magnetic microbeads. Scale bars 500 and 100 nm (inset) (C) Photograph of PDMS iMER cartridge. RT—Reverse transcription (Shao et al., 2015). Source: Reprinted in its original form under CC BY license. License access: https://creativecommons.org/licenses/by/4.0/legalcode.