Seok Hee Lee

Extracellular vesicles (EVs) contain various biological molecules, such as proteins, lipids, and diverse nucleic acids, which alter various physiological and pathological processes in recipient cells. This review focuses on the current understanding of the biological characteristics of EVs on embryo development and their potential therapeutic value in treating reproductive disorders. EVs play a crucial role in early embryo development, from fertilization to the pre-implantation stage, gastrulation, cell differentiation, and organogenesis. During the pre-implantation period, EVs interact with maternal reproductive tissue and promote implantation receptivity. In gastrulation, EVs regulate cell differentiation, contributing to tissue formation and maintenance. Abnormal bioactive molecules in EVs are closely related to developmental disorders. Thus, EVs have the potential to serve as biomarkers. Moreover, EVs can serve as therapeutic agents, delivering genetic material for targeted tissue/organs. The findings of this review highlight the potential role of EVs in intercellular signaling during embryo development. This can help advance assisted reproductive technologies and therapies to overcome infertility issues and developmental disorders.

In vitro fertilization (IVF) has resulted in the birth of over 8 million children. Although most IVF-conceived children are healthy, several studies suggest an increased risk of altered growth rate, cardiovascular dysfunction, and glucose intolerance in this population compared to naturally conceived children. However, a clear understanding of how embryonic metabolism is affected by culture condition and how embryos reprogram their metabolism is unknown. Here, we studied oxidative stress and metabolic alteration in blastocysts conceived by natural mating or by IVF and cultured in physiologic (5%) or atmospheric (20%) oxygen. We found that IVF-generated blastocysts manifest increased reactive oxygen species, oxidative damage to DNA/lipid/proteins, and reduction in glutathione. Metabolic analysis revealed IVF-generated blastocysts display decreased mitochondria respiration and increased glycolytic activity suggestive of enhanced Warburg metabolism. These findings were corroborated by altered intracellular and extracellular pH and increased intracellular lactate levels in IVF-generated embryos. Comprehensive proteomic analysis and targeted immunofluorescence showed reduction of lactate dehydrogenase-B and monocarboxylate transporter 1, enzymes involved in lactate metabolism. Importantly, these enzymes remained downregulated in the tissues of adult IVF-conceived mice, suggesting that metabolic alterations in IVF-generated embryos may result in alteration in lactate metabolism. These findings suggest that alterations in lactate metabolism are a likely mechanism involved in genomic reprogramming and could be involved in the developmental origin of health and disease.

Dogs (Canis lupus familiaris) have unique and peculiar reproductive characteristics. While the interplay between in vitro oviductal cell-derived extracellular vesicles (OC-EVs) and cumulus-oocyte complexes in dogs has begun to be elucidated, no study has yet provided extensive information on the biological content and physiological function of OC-EVs and their role in canine oocyte development. Here, we aimed to provide the first comprehensive proteomic analysis of OC-EVs. We identified 398 proteins as present in all OC-EVs samples. The functional enrichment analysis using Gene Ontology terms and an Ingenuity Pathway Analysis revealed that the identified proteins were involved in several cellular metabolic processes, including translation, synthesis, expression, and protein metabolism. Notably, the proteins were also involved in critical canonical pathways with essential functions in oocyte and embryo development, such as ERK/MAPK, EIF2, PI3K/AKT, and mTOR signaling. These data would be an important resource for studying canine reproductive physiology and establishing a successful in vitro embryo production system in dogs.

An essential requirement for the success of in vitro maturation (IVM) of the oocyte is to provide an optimal microenvironment similar to in vivo conditions. Recently, somatic cell-based coculture or supplementation of a conditioned medium during IVM has been performed to obtain better quality of oocytes, because they mimic the in vivo reproductive tract by secreting paracrine factors. In this study, human adipose-derived stem cells (ASC) and their conditioned medium (ASC-CM) were applied to IVM of porcine oocytes to evaluate the effectiveness of ASC on oocyte development and subsequent embryo development. In results, both ASC and ASC-CM positively influence on oocyte maturation and embryo development by regulating growth factor receptors (VEGF, FGFR, and IGFR), apoptosis (BCL2), cumulus expansion (PTGS2, HAS2, and TNFAIP6), and oocyte maturation-related genes (GDF9 and BMP15). In particular, the fluorescence intensity of GDF9 and BMP15 was markedly upregulated in the oocytes from the ASC-CM group. Furthermore, significantly high levels of growth factors/cytokine including VEGF, bFGF, IGF-1, IL-10, and EGF were observed in ASC-CM. Additionally, the ASC-CM showed active scavenging activity by reducing the ROS production in a culture medium. Consequently, for the first time, this study demonstrated the effect of human ASC-CM on porcine oocyte development and the alteration of mRNA transcript levels in cumulus–oocyte complexes.

Extracellular vesicles (EVs), like exosomes, are nanosized membrane‐enveloped vesicles containing different bioactive cargo, such as proteins, lipids, mRNA, miRNA, and other small regulatory RNAs. Cell‐derived EVs, including EVs originating from stem cells, may capture components from damaged cells or cells impacted by therapeutic treatments. Interestingly, EVs derived from stem cells can be preconditioned to produce and secrete EVs with different therapeutic properties, particularly with respect to heat‐shock proteins and other molecular cargo contents. This behavior is consistent with stem cells that also respond differently to various microenvironments. Heat‐shock proteins play roles in cellular protection and mediate cellular resistance to radiotherapy, chemotherapy, and heat shock. This review highlights the possible roles EVs play in mediating cellular plasticity and survival when exposed to different physical and chemical stressors, with a special focus on the respiratory distress due to the air pollution.

Cryopreservation is an essential tool to preserve sperm cells for zootechnical management and artificial insemination purposes. Cryopreservation is associated with sperm damage via different levels of plasma membrane injury and oxidative stress. Nanoparticles are often used to defend against free radicals and oxidative stress generated through the entire process of cryopreservation. Recently, artificial or natural nanovesicles including liposomes and exosomes, respectively, have shown regenerative capabilities to repair damaged sperm during the freeze–thaw process. Exosomes possess a potential pleiotropic effect because they contain antioxidants, lipids, and other bioactive molecules regulating and repairing spermatozoa. In this review, we highlight the current strategies of using nanoparticles and nanovesicles (liposomes and exosomes) to combat the cryoinjuries associated with semen cryopreservation.

Human endothelial progenitor cells (EPCs) secrete numerous growth factors, and they have been applied to regenerative medicine for their roles in angiogenesis as well as neovascularization. Angiogenesis is one of the essential factors for the maturation of ovarian follicles; however, the physiological function of EPCs or their derivatives on in vitro culture systems has not been fully understood. The aim of this study was to evaluate the effectiveness of EPCs and their conditioned medium (EPC-CM) on oocyte development and subsequent embryo development. In the results, the oocyte development and subsequent embryo development were significantly improved in EPCs and the EPC-CM group. In addition, markedly increased levels of growth factors/cytokines, such as basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), insulin growth factor-1 (IGF-1), interleukin-10 (IL-10), and epidermal growth factor (EGF), were observed in medium from the EPC-CM group. Additionally, EPC-CM after in vitro maturation (IVM) had significantly decreased reactive oxygen species (ROS) levels compared to those of other groups. Transcriptional levels of growth factor receptor-related genes (FGFR2, IGF1R) and anti-apoptotic-related gene (BCL2) were significantly upregulated in cumulus cells/oocytes from the EPC-CM group compared with those from the control. Furthermore, the expression levels of cumulus expansion-related genes (PTGS2, TNFAIP6, HAS2) and oocyte-maturation-related factors (GDF9, BMP15) were significantly enhanced in the EPC-CM group. Consequently, the present study provides the first evidence that EPC-CM contains several essential growth factors for oocyte development by regulating genes involved in oocyte maturation.

The canine oviduct is a unique reproductive organ where the ovulated immature oocytes complete their maturation, while the other mammals ovulate matured gametes. Due to their peculiar reproductive characteristics, the in vitro maturation of dog oocytes is still not wellestablished compared with other mammals. Investigations of the microenvironment conditions in the oviductal canal are required to establish a reliable in vitro maturation system in the dog. Previous studies have suggested that the oviduct and its derivatives play a key role in improving fertilization as well as embryo development. In particular, the biological function of oviduct-derived exosomes on sperm and early embryo development has been investigated in porcine, bovine, and murine species. However, the information about their functions on canine cumulus-oocyte complexes is still elusive. Recent canine reproductive studies demonstrated how oviduct-derived extracellular vesicles such as microvesicles and exosomes interact with oocyte-cumulus complexes and how they can play roles in regulating canine cumulus/oocyte communications. In this review, we summarize the physiological characteristics of canine oviduct-derived exosomes and their potential effects on cumulus cells development as well as oocyte in vitro maturation via molecular signaling pathways.

Lee SH, et al. J Vet Sci. 2019 Oct;21:e7. https://doi.org/10.4142/jvs.2020.21.e7