Exploiting Microfluidics for Extracellular Vesicle Isolation and Characterization: Potential Use for Standardized Embryo Quality Assessment.

Bahram Talebjedi,Marcia De Almeida Monteiro Melo Ferraz,Gabriela F Mastromonaco,Nishat Tasnim,Mina Hoorfar

doi:10.3389/fvets.2020.620809

Abstract

Recent decades have seen a growing interest in the study of extracellular vesicles (EVs), driven by their role in cellular communication, and potential as biomarkers of health and disease. Although it is known that embryos secrete EVs, studies on the importance of embryonic EVs are still very limited. This limitation is due mainly to small sample volumes, with low EV concentrations available for analysis, and to laborious, costly and time-consuming procedures for isolating and evaluating EVs. In this respect, microfluidics technologies represent a promising avenue for optimizing the isolation and characterization of embryonic EVs. Despite significant improvements in microfluidics for EV isolation and characterization, the use of EVs as markers of embryo quality has been held back by two key challenges: (1) the lack of specific biomarkers of embryo quality, and (2) the limited number of studies evaluating the content of embryonic EVs across embryos with varying developmental competence. Our core aim in this review is to identify the critical challenges of EV isolation and to provide seeds for future studies to implement the profiling of embryonic EVs as a diagnostic test for embryo selection. We first summarize the conventional methods for isolating EVs and contrast these with the most promising microfluidics methods. We then discuss current knowledge of embryonic EVs and their potential role as biomarkers of embryo quality. Finally, we identify key ways in which microfluidics technologies could allow researchers to overcome the challenges of embryonic EV isolation and be used as a fast, user-friendly tool for non-invasive embryo selection.

Highlights

The selection of the most fit embryo for transfer after in vitro production remains one of the biggest challenges faced by embryologists today
We aim to summarize the knowledge of the most commonly used methods for extracellular vesicles (EVs) isolation, elaborate on the possibilities offered by microfluidics to improve and standardize current EVs isolation methods, and provide insight on the use of EVs as biomarkers of pre-implantation embryo quality
The combination of these technologies allows the characterization of EV size and concentration, when labeled with lipophilic dyes, and EV DNA cargo if, instead of labeling with lipophilic dyes, EVs are labeled with the DNA marker propidium iodide (PI)

Summary

Introduction

The selection of the most fit embryo (i.e., highest developmental potential to result in a pregnancy and birth of a healthy offspring) for transfer after in vitro production remains one of the biggest challenges faced by embryologists today. Evaluation of embryo quality has been performed by analyzing embryo morphology under a light microscope [1, 2]. It is becoming clear, that embryo competence can be compromised in morphologically normal embryos [3], Microfluidcs for Embryo EVs Characterization as evidenced by differences in in vitro and in vivo embryo transcriptomics and methylomics [4, 5], which could result in implantation failure or miscarriages early in pregnancy [6, 7]. Invasive pre-implantation genomic testing for aneuploidy has improved embryo selection since the transfer of chromosomally normal embryos has resulted in better implantation rates and reduced miscarriages [7, 8]. There lacks a clear understanding of the broader context of the embryonic metabolic environment, and importantly, of which pathways to use to promote optimal quality [15]

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