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Reproduction, Fertility and Embryonic Development of Animals

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A special issue of Animals (ISSN 2076-2615). This special issue belongs to the section "Animal Reproduction".

Deadline for manuscript submissions: closed (28 February 2021) | Viewed by 33799

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Dr. Sandeep K. Rajput

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Colorado Center for Reproductive Medicine (CCRM), Lone Tree, CO 80124, USA
Interests: reproduction; embryonic development; cattle

Special Issue Information

This Special Issue aims to present basic and applied research as well as reviews that increase our understanding of reproductive processes, fertility issues and treatments, and early embryonic development in small and large animals. Studies focusing on reproductive technologies, physiological and molecular mechanisms, and applied genetics to increase fertility in livestock animals and conservation of rare species are of particular interest.

The scope of this Special Issue includes reproductive health; endocrinology; infertility; early embryo development, pregnancy; the development of molecular markers to assess fertility, sperm/oocyte/embryo quality, and reproductive disorders as well as to design breeding strategies; assisted reproductive technologies such as vitrification, intracytoplasmic sperm injection (ICSI), in vitro fertilization (IVF), genetic analysis by embryo biopsies, embryo transfer, animal cloning, fertility preservation, and artificial insemination; molecular and metabolic understanding of oocyte and embryo development; oocyte and embryo culture; embryo outgrowth; and computer-assisted semen and embryo analysis. We especially welcome manuscripts that highlight the use of the latest technologies in addressing problems related to reproduction, fertility, and early embryonic development, such as CRISPR–Cas9 and lentivirus-mediated genomic manipulation, single-cell RNA sequencing, and proteomic analysis. We hope that this issue will provide novel insights into advancements in the field of reproduction and fertility in livestock and wild animals.

Dr. Sandeep K. Rajput
Guest Editor

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Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All submissions that pass pre-check are peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. Animals is an international peer-reviewed open access semimonthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. The Article Processing Charge (APC) for publication in this open access journal is 2400 CHF (Swiss Francs). Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Keywords

animal
oocytes
embryos
fertility
pregnancy
genetics
assisted reproductive technologies
metabolism
next-generation sequencing
genomic manipulation

Published Papers (11 papers)

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Research

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11 pages, 2155 KiB

Open AccessArticle

V-Set and Immunoglobulin Domain-Containing 1 (VSIG1), Predominantly Expressed in Testicular Germ Cells, Is Dispensable for Spermatogenesis and Male Fertility in Mice

by Yena Jung, Hyewon Bang, Young-Hyun Kim, Na-Eun Park, Young-Ho Park, Chaeli Park, Sang-Rae Lee, Jeong-Woong Lee, Bong-Seok Song, Ji-Su Kim, Bo-Woong Sim, Dong-Won Seol, Gabbine Wee, Sunhyung Kim, Sun-Uk Kim and Ekyune Kim

Animals 2021, 11(4), 1037; https://0-doi-org.brum.beds.ac.uk/10.3390/ani11041037 - 07 Apr 2021

Cited by 5 | Viewed by 2320

Abstract

To elucidate the functional role of V-set and immunoglobulin domain-containing 1 (VSIG1) in spermatogenesis and fertilization, we knocked out (KO) VSIG1 in a mouse embryo using CRISPR/Cas9 (Clustered regularly interspaced short palindromic repeat/CRISPR-associated protein 9) -mediated genome editing. Reverse transcription PCR was performed using cDNA synthesized from VSIG1 KO testis RNA. Although Western blot analysis using a specific antibody to VSIG1 confirmed VSIG1 protein defects in the KO mice, hematoxylin-eosin staining analysis was similar in the KO and wild-type mice. Additionally, computer-assisted sperm analysis and in vitro fertilization experiments were conducted to confirm the activity and fertilization ability of sperm derived from the KO mouse. Mice lacking VSIG1 were viable and had no serious developmental defects. As they got older, the KO mice showed slightly higher weight loss, male mice lacking VSIG1 had functional testes, including normal sperm number and motility, and both male and female mice lacking VSIG1 were fertile. Our results from VSIG1 KO mice suggest that VSIG1 may not play essential roles in spermatogenesis and normal testis development, function, and maintenance. VSIG1 in sperm is dispensable for spermatogenesis and male fertility in mice. As several genes are known to possess slightly different functions depending on the species, the importance and molecular mechanism of VSIG1 in tissues of other species needs further investigation. Full article

(This article belongs to the Special Issue Reproduction, Fertility and Embryonic Development of Animals)

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14 pages, 1753 KiB

Open AccessArticle

Effect of D-Glucuronic Acid and N-acetyl-D-Glucosamine Treatment during In Vitro Maturation on Embryonic Development after Parthenogenesis and Somatic Cell Nuclear Transfer in Pigs

by Joohyeong Lee, Eunhye Kim, Seon-Ung Hwang, Lian Cai, Mirae Kim, Hyerin Choi, Dongjin Oh, Eunsong Lee and Sang-Hwan Hyun

Animals 2021, 11(4), 1034; https://0-doi-org.brum.beds.ac.uk/10.3390/ani11041034 - 06 Apr 2021

Cited by 5 | Viewed by 2585

Abstract

This study aimed to examine the effects of treatment with glucuronic acid (GA) and N-acetyl-D-glucosamine (AG), which are components of hyaluronic acid (HA), during porcine oocyte in vitro maturation (IVM). We measured the diameter of the oocyte, the thickness of the perivitelline space (PVS), the reactive oxygen species (ROS) level, and the expression of cumulus cell expansion and ROS-related genes and examined the cortical granule (CG) reaction of oocytes. The addition of 0.05 mM GA and 0.05 mM AG during the first 22 h of oocyte IVM significantly increased oocyte diameter and PVS size compared with the control (non-treatment). The addition of GA and AG reduced the intra-oocyte ROS content and improved the CG of the oocyte. GA and AG treatment increased the expression of CD44 and CX43 in cumulus cells and PRDX1 and TXN2 in oocytes. In both the chemically defined and the complex medium (Medium-199 + porcine follicular fluid), oocytes derived from the GA and AG treatments presented significantly higher blastocyst rates than the control after parthenogenesis (PA) and somatic cell nuclear transfer (SCNT). In conclusion, the addition of GA and AG during IVM in pig oocytes has beneficial effects on oocyte IVM and early embryonic development after PA and SCNT. Full article

(This article belongs to the Special Issue Reproduction, Fertility and Embryonic Development of Animals)

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14 pages, 1976 KiB

Open AccessArticle

Effect of Interleukin-7 on In Vitro Maturation of Porcine Cumulus-Oocyte Complexes and Subsequent Developmental Potential after Parthenogenetic Activation

by Dongjin Oh, Joohyeong Lee, Eunhye Kim, Seon-Ung Hwang, Junchul-David Yoon, Lian Cai, Mirae Kim, Gahye Kim, Hyerin Choi and Sang-Hwan Hyun

Animals 2021, 11(3), 741; https://0-doi-org.brum.beds.ac.uk/10.3390/ani11030741 - 08 Mar 2021

Cited by 7 | Viewed by 2776

Abstract

Interleukin-7 (IL-7) is a cytokine essential for cell development, proliferation and survival. However, its role in oocyte maturation is largely unknown. To investigate the effects of IL-7 on the in vitro maturation (IVM) of porcine oocytes, we analyzed nuclear maturation, intracellular glutathione (GSH) and reactive oxygen species (ROS) levels, and subsequent embryonic developmental competence after parthenogenetic activation (PA) under several concentrations of IL-7. After IVM, IL-7 treated groups showed significantly higher nuclear maturation and significantly decreased intracellular ROS levels compared with the control group. All IL-7 treatment groups exhibited significantly increased intracellular GSH levels compared with the control group. All oocytes matured with IL-7 treatment during IVM exhibited significantly higher cleavage and blastocyst formation rates after PA than the non-treatment group. Furthermore, significantly higher mRNA expression levels of developmental-related genes (PCNA, Filia, and NPM2) and antioxidant-related genes (GSR and PRDX1) were observed in the IL-7-supplemented oocytes than in the control group. IL-7-supplemented cumulus cells showed significantly higher mRNA expression of the anti-apoptotic gene BCL2L1 and mitochondria-related genes (TFAM and NOX4), and lower transcript levels of the apoptosis related-gene, Caspase3, than the control group. Collectively, the present study suggests that IL-7 supplementation during porcine IVM improves oocyte maturation and the developmental potential of porcine embryos after PA. Full article

(This article belongs to the Special Issue Reproduction, Fertility and Embryonic Development of Animals)

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17 pages, 23271 KiB

Open AccessArticle

R-Spondin 2 and WNT/CTNNB1 Signaling Pathways Are Required for Porcine Follicle Development and In Vitro Maturation

by Seon-Ung Hwang, Junchul David Yoon, Mirae Kim, Lian Cai, Hyerin Choi, Dongjin Oh, Eunhye Kim and Sang-Hwan Hyun

Animals 2021, 11(3), 709; https://0-doi-org.brum.beds.ac.uk/10.3390/ani11030709 - 05 Mar 2021

Cited by 8 | Viewed by 2187

Abstract

The secretion of oocyte-derived paracrine factors, such as R-spondin2, is an essential mechanism for follicle growth by promoting the proliferation and differentiation of cumulus cells around oocytes. In the present study, we aimed to identify the effect of R-spondin2 during follicular development. First, R-spondin2-related factors (R-spondin2, CTNNB1, LGR4, and LGR5) were identified through immunofluorescence in porcine ovarian tissue. CTNNB1 was expressed in ooplasm, and CTNNB1 and LGR4 were expressed in granulosa cells. In addition, R-spondin2, LGR4, and LGR5 were expressed in the theca interna. These results imply that these proteins play a major role in porcine follicular development. In addition, the effects of R-spondin2 on the in vitro maturation process of porcine cumulus oocyte complexes and subsequent embryonic development were confirmed. A treatment of 100 ng/mL R-spondin2 in the in vitro maturation (IVM) process increased nuclear maturation and increased the expression of EGFR mRNA in cumulus cells. The EGFR-ERK signal is essential for oocyte maturation, ovulation, and luteinization. R-spondin2 treatment also increased the expression of CTNNB1 and EGFR in primary cultured cumulus cells. In conclusion, RSPO2 and WNT/CTNNB1 signaling pathways are required for porcine follicle development and are predicted to be involved in the EGFR-ERK signaling pathway. Full article

(This article belongs to the Special Issue Reproduction, Fertility and Embryonic Development of Animals)

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16 pages, 2029 KiB

Open AccessArticle

Adiponectin Improves In Vitro Development of Cloned Porcine Embryos by Reducing Endoplasmic Reticulum Stress and Apoptosis

by Muhammad Rosyid Ridlo, Eui Hyun Kim, Anukul Taweechaipaisankul, Byeong Chun Lee and Geon A. Kim

Animals 2021, 11(2), 473; https://0-doi-org.brum.beds.ac.uk/10.3390/ani11020473 - 10 Feb 2021

Cited by 4 | Viewed by 1940

Abstract

The main factor of embryonic demise is endoplasmic reticulum (ER) stress. Successful attenuation of ER stress results in an improvement in embryo development. We studied the impact of adiponectin in the in vitro culture (IVC) of porcine embryos derived from parthenogenetic activation and somatic cell nuclear transfer (SCNT). The first experiment revealed that 15 and 30 μg/mL adiponectin treatments improved cleavage, blastocyst rates, and total cell number (TCN) of parthenogenetic embryos and reduced the expression of XBP1 compared to the 5 μg/mL adiponectin treatment and control groups (p < 0.05). The second experiment showed that cleavage rate, blastocyst formation rate, and TCN of blastocysts were improved in the 15 μg/mL adiponectin treatment group compared with the control group, with significantly reduced XBP1 expression in ≥4-cell stage SCNT embryos and blastocysts (p < 0.05). Treatment with 15 μg/mL adiponectin significantly improved the expression of XBP1 and reduced the expression of ER stress-related genes (uXBP1, sXBP1, PTPN1, and ATF4), increased the expression levels of pluripotency-related genes (Nanog and SOX2), and decreased apoptosis-related gene expression (Caspase-3). These results suggest that 15 μg/mL adiponectin enhanced the in vitro developmental capacity of early-stage SCNT porcine embryos by reducing ER stress and apoptosis. Full article

(This article belongs to the Special Issue Reproduction, Fertility and Embryonic Development of Animals)

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12 pages, 1474 KiB

Open AccessArticle

Inhibition of miR-152 during In Vitro Maturation Enhances the Developmental Potential of Porcine Embryos

by Ahmed Gad, Matej Murin, Lucie Nemcova, Alexandra Bartkova, Jozef Laurincik and Radek Procházka

Animals 2020, 10(12), 2289; https://0-doi-org.brum.beds.ac.uk/10.3390/ani10122289 - 04 Dec 2020

Cited by 3 | Viewed by 2353

Abstract

Oocyte developmental competence is regulated by various mechanisms and molecules including microRNAs (miRNAs). However, the functions of many of these miRNAs in oocyte and embryo development are still unclear. In this study, we managed to manipulate the expression level of miR-152 during oocyte maturation to figure out its potential role in determining the developmental competence of porcine oocytes. The inhibition (Inh) of miR-152 during oocyte maturation does not affect the MII and cleavage rates, however it significantly enhances the blastocyst rate compared to the overexpression (OvExp) and control groups. Pathway analysis identified several signaling pathways (including PI3K/AKT, TGFβ, Hippo, FoxO, and Wnt signaling) that are enriched in the predicted target genes of miR-152. Gene expression analysis revealed that IGF1 was significantly up-regulated in the Inh group and downregulated in the OvExp group of oocytes. Moreover, IGF1R was significantly upregulated in the Inh oocyte group compared to the control one and IGFBP6 was downregulated in the Inh oocyte group compared to the other groups. Blastocysts developed from the OvExp oocytes exhibited an increase in miR-152 expression, dysregulation in some quality-related genes, and the lowest rate of blastocyst formation. In conclusion, our results demonstrate a negative correlation between miR-152 expression level and blastocyst rate in pigs. This correlation could be through targeting IGF system components during oocyte development. Full article

(This article belongs to the Special Issue Reproduction, Fertility and Embryonic Development of Animals)

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13 pages, 639 KiB

Open AccessArticle

Meiotic Status Does Not Affect the Vitrification Effectiveness of Domestic Cat Oocytes

by Natalia Sowińska, Jennifer Zahmel, Wojciech Niżański, Romy Hribal, Lorena Fernandez-Gonzalez and Katarina Jewgenow

Animals 2020, 10(8), 1371; https://0-doi-org.brum.beds.ac.uk/10.3390/ani10081371 - 07 Aug 2020

Cited by 9 | Viewed by 2679

Abstract

Cryopreservation is important for animal fertility and biodiversity. Unfortunately, cryopreservation of feline oocytes is still an experimental technique. The aims of this study were to analyze the potential toxicity of the cryoprotectants in the vitrification solution (VS) on cat oocytes and to investigate whether the meiotic status of oocytes influences their developmental potential after vitrification. Two experiments were conducted with the VS composed of 20% ethylene glycol, 20% dimethyl sulfoxide, 20% fetal calf serum, 1.5 M trehalose, and 10% Ficoll PM-70: (1) toxicity assessment of the VS on immature cumulus oocyte complexes (COCs), and subsequently in vitro maturation (IVM) and in vitro fertilization; (2) assessment of the influence of the meiotic status on vitrification effectiveness, where immature and in vitro matured COCs were vitrified on the Cryotop. After rewarming, vitrified oocytes were subjected to IVM (immature) and intracytoplasmic sperm injection (ICSI) with fresh epididymal sperm. The toxicity test revealed no negative effect of oocyte exposure to the applied VS on their developmental potential (p > 0.05). Although the vitrification procedure itself significantly reduced the meiotic competence of oocytes, their meiotic status before vitrification (immature vs. in vitro matured) did not influence fertilization and morula rates. The only parameter affected by vitrification was the rate of oocytes suitable for ICSI, which was significantly lower for immature oocytes. Regardless of the meiotic status of vitrified oocytes, morphologically normal morulae were obtained. Moreover, the two meiotic stages examined are suitable for vitrification, with mature oocytes being a better choice when a well-equipped laboratory is available. Full article

(This article belongs to the Special Issue Reproduction, Fertility and Embryonic Development of Animals)

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15 pages, 4712 KiB

Open AccessArticle

Endometrial Cytology During the Different Phases of the Estrous Cycle in Jennies: New Evidences

by Marco Quartuccio, Santo Cristarella, Pietro Medica, Esterina Fazio, Giuseppe Mazzullo, Claudia Rifici, Luigi Liotta and Katiuska Satué

Animals 2020, 10(6), 1062; https://0-doi-org.brum.beds.ac.uk/10.3390/ani10061062 - 19 Jun 2020

Cited by 4 | Viewed by 6682

Abstract

Since in the mare and other animal species such as bitches and cats, the endometrial cell pattern varies depending on the phase of the estrous cycle, the aim of this study was to describe and quantify the endometrial cytological (EC) findings in cycling jennies. EC of eight nonpregnant jennies by cytobrush (CB) at diestrus (day 1 and day 14) and estrous (day 21) were evaluated. All slides were stained with Wright´s stain and microscopically examined at both 400× and 1000× magnification. Seven high-power fields (400×) were assessed in each smear and the endometrial epithelial cells and neutrophils (PMNs) were counted. Endometrial epithelial cells were classified as intact, distorted or fragmented and, on the basis of the presence of dense groups, in monolayer or single clusters. Cytoplasmic characteristics, such as vacuolation or streaming and size, form, position of nuclear characteristics, including karyorrhexis, were recorded. Background aspect, as clear, proteinaceous, or debris, was also considered. In general, sampling by CB provided a yield of cells and clumped endometrial epithelial cells in many smears, being more abundant in estrus than early and late diestrus. Individual endometrial epithelial cells, during estrous, presented a columnar morphology, ciliated or not ciliated and basal nuclei. During diestrus phase, endometrial epithelial cells presented a more cuboidal ciliated or not ciliated morphology. Moderate amount of proteinacious material and red blood cells (RBC) was also observed. Non variation in the percentage of PMNs during diestrus was obtained, but lower and segmented PMNs in CB smears were shown in estrous. This study provides new insights on the physiological changes of endometrial epithelial cells in cycling jennies during the estrus cycle. The CB technique represents a suitable and adequate method for endometrial evaluation, taking into account cytological and/or cytopathological purposes also in jennies. Full article

(This article belongs to the Special Issue Reproduction, Fertility and Embryonic Development of Animals)

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14 pages, 3401 KiB

Open AccessArticle

High Frequency of Intravenous Injection of Human Adipose Stem Cell Conditioned Medium Improved Embryo Development of Mice in Advanced Maternal Age through Antioxidant Effects

by Kihae Ra, Hyun Ju Oh, Geon A Kim, Sung Keun Kang, Jeong Chan Ra and Byeong Chun Lee

Animals 2020, 10(6), 978; https://0-doi-org.brum.beds.ac.uk/10.3390/ani10060978 - 04 Jun 2020

Cited by 8 | Viewed by 2275

Abstract

Advanced maternal age (AMA) has become prevalent globally. With aging, weakened antioxidant defense causes loss of normal function in the ovary and uterus due to oxidative stress. Here, we aimed to improve embryo development in AMA mice by intravenous injection (IV) of human adipose stem cell conditioned medium (ASC-CM) at various frequencies and intervals as an antioxidant intervention. Four- and six-month-old female ICR (Institute of Cancer Research) mice were randomly divided into groups IV treated with human ASC-CM under different conditions, and in vitro and in vivo embryo development were evaluated. Consequently, compared to the control group, blastocyst formation rate of parthenotes was significantly promoted in 4-month-old mice and the mean number of implanted fetuses after natural mating was significantly increased by approximately two-fold in 6-month-old mice. Through gene analysis, the anti-apoptotic and anti-oxidative effects of human ASC-CMs were confirmed in the ovaries and uterus of pregnant mice at both ages. In particular, ovarian expression of gpx1 and catalase drastically increased in 6-month-old mice. Furthermore, the levels of gpx1 and catalase were further increased, with a high frequency of injection regardless of age. Thus, we demonstrated for the first time the anti-oxidative effect of human ASC-CM administration against ovarian aging and the optimal injection condition. Full article

(This article belongs to the Special Issue Reproduction, Fertility and Embryonic Development of Animals)

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17 pages, 1731 KiB

Open AccessArticle

Developmental Plasticity in Response to Embryo Cryopreservation: The Importance of the Vitrification Device in Rabbits

by Ximo Garcia-Dominguez, José Salvador Vicente and Francisco Marco-Jiménez

Animals 2020, 10(5), 804; https://0-doi-org.brum.beds.ac.uk/10.3390/ani10050804 - 06 May 2020

Cited by 15 | Viewed by 3095

Abstract

In this study, we evaluated the effect of embryo vitrification using two different devices on adulthood phenotype in rabbits. In vitro development, prenatal embryo survival, body weight, growth performance, haematological and biochemical peripheral blood analysis, reproductive performance, and lactation performance traits were compared between the experimental groups. They derived from naturally-conceived embryos (NC), fresh-transferred embryos (FT), vitrified-transferred embryos using mini-straw (VTs), or vitrified-transferred embryos using Cryotop (VTc). Straw-vitrified embryos exhibited lower in vitro developmental rates and in vivo survival rates following embryo transfer compared to its Cryotop-vitrified counterparts. Moreover, the VTs group exhibited higher foetal losses than VTc, FT, and NC groups. Independently of the vitrification device, vitrified-transferred (VT) offspring showed a skewed sex ratio in favour of males, and an increased birth bodyweight. In contrast, postnatal daily growth was diminished in all ART (i.e., FT and VT) animals. In adulthood, significant differences in body weight between all groups was founded—all ART progenies weighed less than NC animals and, within ART, VT animals weighed less than FT. For VT groups, weight at adulthood was higher for the VTs group compared with the VTc group. Peripheral blood parameters ranged between common values. Moreover, no differences were found in the fertility rates between experimental groups. Furthermore, similar pregnancy rates, litter sizes, and the number of liveborns were observed, regardless of the experimental group. However, decreased milk yield occurred for VTc and FT animals compared to VTs and NC animals. A similar trend was observed for the milk composition of dry matter and fat. Concordantly, reduced body weight was found for suckling kits in the VTc and FT groups compared to VTs and NC animals. Our findings reveal that developmental changes after the embryo vitrification procedure could be associated with an exhibition of the embryonic developmental plasticity. Moreover, to our best knowledge, this study reports the first evidence demonstrating that the vitrification device used is not a trivial decision, providing valuable information about how the cooling–warming rates during vitrification can be partly responsible of the postnatal phenotypic variations. Full article

(This article belongs to the Special Issue Reproduction, Fertility and Embryonic Development of Animals)

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Review

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19 pages, 397 KiB

Open AccessReview

Overview on the Antioxidants, Egg Yolk Alternatives, and Mesenchymal Stem Cells and Derivatives Used in Canine Sperm Cryopreservation

by Feriel Yasmine Mahiddine and Min-Jung Kim

Animals 2021, 11(7), 1930; https://0-doi-org.brum.beds.ac.uk/10.3390/ani11071930 - 28 Jun 2021

Cited by 13 | Viewed by 3029

Abstract

Sperm cryopreservation is a widely used assisted reproductive technology for canine species. The long-term storage of dog sperm is effective for the breeding of dogs living far apart, scheduling the time of artificial insemination that suits the female, and preventing diseases of the reproductive tract. However, spermatozoa functions are impaired during the freeze–thaw processes, which may decrease reproductive performance. Numerous attempts have been made to restore such impairments, including the use of cryoprotectants to prevent the damage caused by ice crystal formation, and supplementation of antioxidants to reduce reactive oxygen species generation due to osmotic stress during the procedure. Egg yolk derivatives, antioxidants, and, more recently, mesenchymal stem cells (MSCs) and their derivatives have been proposed in this research field. This review article will summarize the current literature available on the topic. Full article

(This article belongs to the Special Issue Reproduction, Fertility and Embryonic Development of Animals)

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