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Article ; Online: Synchronization of the ovulation and copulation timings increased the number of in vivo fertilized oocytes in superovulated female mice.

Satohiro Nakao / Kotono Ito / Chihiro Sugahara / Hitomi Watanabe / Gen Kondoh / Naomi Nakagata / Toru Takeo

PLoS ONE, Vol 18, Iss 2, p e

2023 Volume 0281330

Abstract: The number of sperm that reaches the oocytes in mammalian species is limited. In mice, 8-10 oocytes are ovulated, a similar number of sperm reaches the oocytes, and nearly all oocytes are fertilized via natural mating. Meanwhile, our improved ... ...

Abstract	The number of sperm that reaches the oocytes in mammalian species is limited. In mice, 8-10 oocytes are ovulated, a similar number of sperm reaches the oocytes, and nearly all oocytes are fertilized via natural mating. Meanwhile, our improved superovulation technique (ultrasuperovulation: administration of inhibin antiserum and equine chorionic gonadotropin [IASe]) produced 100 oocytes from a single female C57BL/6 mouse but resulted in only approximately 20 fertilized oocytes via mating. We hypothesized that sperm shortage in the ampulla might cause this low fertilization rate. Mice were mated in the proestrus stage or after hormone injection, but ovulation timing was not considered. In clinical application, the rhythm method supports fertilization by testing the ovulation period and synchronizing the ovulation and copulation timings. Therefore, this study examined the effects of ovulation and copulation timings on in vivo fertilization in female mice with IASe. Synchronization of the ovulation and copulation timings increased fertilization efficiency in female mice with ultrasuperovulation. The number of embryos obtained post ovulation was three times higher than that obtained pre ovulation. This study suggests that synchronized ovulation and copulation timings improve the efficiency of in vivo fertilization in IASe-treated female mice. This technique can be used to produce genetically modified mice and develop technologies for infertility treatment.
Keywords	Medicine ; R ; Science ; Q
Subject code	630
Language	English
Publishing date	2023-01-01T00:00:00Z
Publisher	Public Library of Science (PLoS)
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: N-acetyl cysteine restores the fertility of vitrified-warmed mouse oocytes derived through ultrasuperovulation.

Ayumi Mukunoki / Toru Takeo / Naomi Nakagata

PLoS ONE, Vol 14, Iss 10, p e

2019 Volume 0224087

Abstract: Oocyte cryopreservation is useful for preserving fertility and storing genetic resources. However, the small number of oocytes acquired using conventional treatment to induce superovulation and the reduction of fertility due to cryopreservation represent ...

Abstract	Oocyte cryopreservation is useful for preserving fertility and storing genetic resources. However, the small number of oocytes acquired using conventional treatment to induce superovulation and the reduction of fertility due to cryopreservation represent significant problems. Herein, we vitrified the oocytes derived through high-yield superovulation using inhibin antiserum and equine chorionic gonadotropin (IAS + eCG: IASe) and examined the yield of cryopreserved oocytes and survival rates relative to those of vitrified-warmed mouse oocytes derived through conventional superovulation using equine chorionic gonadotropin (eCG). Furthermore, we investigated the effects of N-acetyl cysteine on the fertility and developmental potential of vitrified-warmed oocytes derived using IASe. Compared with eCG, IASe increased the yield of cryopreserved oocytes and achieved equivalent survival rates. N-acetyl cysteine (0.5 mM) increased the fertilization rate of vitrified-warmed oocytes derived using IASe. Vitrification decreased thiol levels in the zona pellucida (ZP), while warming followed by N-acetyl cysteine treatment increased free thiol levels in ZP. Moreover, N-acetyl cysteine treatment recovered zona hardening by cleaving disulfide bonds and promoting the expansion of ZP. Two-cell embryos derived via in vitro fertilization using N-acetyl cysteine developed into normal pups through embryo transfer. Therefore, we developed an efficient technique for the production of cryopreserved oocytes using IASe through superovulation and found that N-acetyl cysteine improves the fertility of vitrified-warmed oocytes by cleaving the disulfide bonds and promoting the expansion of ZP.
Keywords	Medicine ; R ; Science ; Q
Language	English
Publishing date	2019-01-01T00:00:00Z
Publisher	Public Library of Science (PLoS)
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: Establishment of sperm cryopreservation and in vitro fertilisation protocols for rats

Naomi Nakagata / Nobuyuki Mikoda / Satohiro Nakao / Ena Nakatsukasa / Toru Takeo

Scientific Reports, Vol 10, Iss 1, Pp 1-

2020 Volume 8

Abstract: Abstract Recently, genome-editing tools have come into common use in the field of rat research, and consequently, many genetically modified rat strains have been preserved and archived as frozen embryos. Although there have been many reports published on ...

Abstract	Abstract Recently, genome-editing tools have come into common use in the field of rat research, and consequently, many genetically modified rat strains have been preserved and archived as frozen embryos. Although there have been many reports published on the topic of rat sperm cryopreservation, no report has yet provided satisfactory and acceptable protocols for the cryopreservation of rat sperm. In this study, we developed methods for both the cryopreservation of transgenic rat sperm and in vitro fertilisation using frozen sperm, which yielded high fertilisation rates.
Keywords	Medicine ; R ; Science ; Q
Language	English
Publishing date	2020-01-01T00:00:00Z
Publisher	Nature Publishing Group
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: Successful selection of mouse sperm with high viability and fertility using microfluidics chip cell sorter

Satohiro Nakao / Toru Takeo / Hitomi Watanabe / Gen Kondoh / Naomi Nakagata

Scientific Reports, Vol 10, Iss 1, Pp 1-

2020 Volume 9

Abstract: Abstract Cell sorting via flow cytometry is a powerful tool to select subpopulations of cells in many biological fields. Selection of fertilisation-prone sperm is a critical step to ensure a stable and high fertilisation rate in in vitro fertilisation ( ... ...

Abstract	Abstract Cell sorting via flow cytometry is a powerful tool to select subpopulations of cells in many biological fields. Selection of fertilisation-prone sperm is a critical step to ensure a stable and high fertilisation rate in in vitro fertilisation (IVF). However, a combination of conventional cell sorting and IVF system has not been established because of severe mechanical damages to the sperm during the sorting process. A cell sorter with microfluidics chip technology that lessens cell damage during cell sorting may address this problem. We evaluated the effects of microfluidics chip cell sorting on the sperm using the parameters, such as motility and fertility, and found this cell sorting method had minimal harmful effect on the sperm. Then, sperm were selected by a marker for acrosome reaction and showed higher fertilisation rate than that of the population of acrosome-intact sperm. Embryo derived from these sperm developed normally. These results indicated that microfluidics chip cell sorting can select fertile sperm to improve IVF technique.
Keywords	Medicine ; R ; Science ; Q
Subject code	571
Language	English
Publishing date	2020-06-01T00:00:00Z
Publisher	Nature Publishing Group
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: Quercetin-treated rat sperm enables refrigerated transport with motility and fertility for five days

Katsuma Yamaga / Satohiro Nakao / Nobuyuki Mikoda / Hidetaka Yoshimoto / Ena Nakatsukasa / Naomi Nakagata / Toru Takeo

Scientific Reports, Vol 11, Iss 1, Pp 1-

2021 Volume 9

Abstract: Abstract Shipment of laboratory rats between animal facilities is frequently performed using special containers. However, the shipment of live animals is associated with potential risks of infectious diseases, escape and death during shipment and animal ... ...

Abstract	Abstract Shipment of laboratory rats between animal facilities is frequently performed using special containers. However, the shipment of live animals is associated with potential risks of infectious diseases, escape and death during shipment and animal welfare issues. The transport of cold-stored sperm avoids such risks; however, there have been no reports on cold storage of rat sperm. We previously reported that dimethyl sulfoxide (DMSO) and quercetin maintained the motility and fertilising abilities of cold-stored mouse sperm stored for 10 days. The present study investigated the efficacy of DMSO and quercetin in the cold storage of rat sperm. Quercetin maintained motility and fertility of cold-stored rat sperm stored for 5 days. After in vitro fertilisation using cold-stored sperm, pronuclear and two-cell embryos developed normally to pups following embryo transfer. Therefore, we demonstrated that live pups could be obtained from sperm transported using the cold-storage system. We conclude that cold storage of rat sperm may provide an efficient system for transporting rat resources as an alternative to shipping live animals.
Keywords	Medicine ; R ; Science ; Q
Subject code	630
Language	English
Publishing date	2021-11-01T00:00:00Z
Publisher	Nature Portfolio
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: Superovulation using the combined administration of inhibin antiserum and equine chorionic gonadotropin increases the number of ovulated oocytes in C57BL/6 female mice.

Toru Takeo / Naomi Nakagata

PLoS ONE, Vol 10, Iss 5, p e

2015 Volume 0128330

Abstract: Superovulation is a reproductive technique generally used to produce genetically engineered mice. Superovulation in mice involves the administration of equine chorionic gonadotropin (eCG) to promote follicle growth and then that of human chorionic ... ...

Abstract	Superovulation is a reproductive technique generally used to produce genetically engineered mice. Superovulation in mice involves the administration of equine chorionic gonadotropin (eCG) to promote follicle growth and then that of human chorionic gonadotropin (hCG) to induce ovulation. Previously, some published studies reported that inhibin antiserum (IAS) increased the number of ovulated oocytes in ddY and wild-derived strains of mice. However, the effect of IAS on the C57BL/6 strain, which is the most widely used inbred strain for the production of genetically engineered mice, has not been investigated. In addition, the combined effect of IAS and eCG (IASe) on the number of ovulated oocytes in superovulation treatment has not been examined. In this study, we examined the effect of IAS and eCG on the number of ovulated oocytes in immature female mice of the C57BL/6 strain in superovulation treatment. Furthermore, we evaluated the quality of obtained oocytes produced by superovulation using IASe by in vitro fertilization (IVF) with sperm from C57BL/6 or genetically engineered mice. The developmental ability of fresh or cryopreserved embryos was examined by embryo transfer. The administration of IAS or eCG had a similar effect on the number of ovulated oocytes in C57BL/6 female mice. The number of ovulated oocytes increased to about 3-fold by the administration of IASe than by the administration of IAS or eCG alone. Oocytes derived from superovulation using IASe normally developed into 2-cell embryos by IVF using sperm from C57BL/6 mice. Fresh or cryopreserved 2-cell embryos produced by IVF between oocytes of C57BL/6 mice and sperm from genetically engineered mice normally developed into live pups following embryo transfer. In summary, a novel technique of superovulation using IASe is extremely useful for producing a great number of oocytes and offspring from genetically engineered mice.
Keywords	Medicine ; R ; Science ; Q
Subject code	630
Language	English
Publishing date	2015-01-01T00:00:00Z
Publisher	Public Library of Science (PLoS)
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: Cryopreservation of mouse resources

Toru Takeo / Satohiro Nakao / Yoshiko Nakagawa / Jorge M. Sztein / Naomi Nakagata

Laboratory Animal Research, Vol 36, Iss 1, Pp 1-

2020 Volume 6

Abstract: Abstract The cryopreservation of sperm and embryos is useful to efficiently archive valuable resources of genetically engineered mice. Till date, more than 60,000 strains of genetically engineered mice have been archived in mouse banks worldwide. ... ...

Abstract	Abstract The cryopreservation of sperm and embryos is useful to efficiently archive valuable resources of genetically engineered mice. Till date, more than 60,000 strains of genetically engineered mice have been archived in mouse banks worldwide. Researchers can request for the archived mouse strains for their research projects. The research infrastructure of mouse banks improves the availability of mouse resources, the productivity of research projects, and the reproducibility of animal experiments. Our research team manages the mouse bank at the Center for Animal Resources and Development in Kumamoto University and continuously develops new techniques in mouse reproductive technology to efficiently improve the system of mouse banking. In this review, we introduce the activities of mouse banks and the latest techniques used in mouse reproductive technology.
Keywords	Genetically engineered mice ; Mouse bank ; Reproductive technology ; Sperm ; Embryo ; Cryopreservation ; Medicine (General) ; R5-920 ; Biology (General) ; QH301-705.5
Subject code	027
Language	English
Publishing date	2020-09-01T00:00:00Z
Publisher	BMC
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article: Immunotherapy using inhibin antiserum enhanced the efficacy of equine chorionic gonadotropin on superovulation in major inbred and outbred mice strains

Takeo, Toru / Naomi Nakagata

Theriogenology. 2016 Sept. 15, v. 86, no. 5

2016

Abstract: Improvement of the superovulation technique will help to enhance the efficiency of embryo and animal production. Blocking inhibin using inhibin antiserum (IAS) is known to promote follicular development by increasing the level of FSH. Previously, we ... ...

Abstract	Improvement of the superovulation technique will help to enhance the efficiency of embryo and animal production. Blocking inhibin using inhibin antiserum (IAS) is known to promote follicular development by increasing the level of FSH. Previously, we reported that coadministration of IAS and eCG produced more than 100 oocytes from a single female C57BL/6 mouse at 4 weeks old. The oocytes derived from the IAS + eCG (IASe) treatment were able to fertilize and develop normally into offspring. In this study, we examined the effect of IASe treatment on the numbers of ovulated oocytes in major inbred (A/J, BALB/cByJ, C3HeJ, DBA/2J, and FVB/NJ) and outbred (CD1) mice strains at 4 weeks old. We confirmed the fertilization and developmental ability of the IASe-derived oocytes. IASe treatment ovulated 1.5 to 3.2 times higher numbers of oocytes than eCG treatment alone. The fertilization rate of IASe-derived oocytes was similar to that of eCG-derived oocytes. In vitro and in vivo developmental rates of the embryos derived from IASe were similar to the rates of embryos derived from eCG. We have shown that superovulation by IASe is very effective in obtaining high numbers of ovulated oocytes from small numbers of oocyte donor in a number of mice strains. The superovulation technique will contribute to the archiving of cryopreserved embryos of genetically engineered mice using small numbers of donors and has the potential to produce more live animals for rederivation of the archived mouse lines in mouse repositories.
Keywords	animal production ; antiserum ; cryopreservation ; embryo (animal) ; equine chorionic gonadotropin ; females ; fertilizer rates ; follicle-stimulating hormone ; follicular development ; genetic engineering ; immunotherapy ; inhibin ; mice ; oocytes ; progeny ; superovulation
Language	English
Dates of publication	2016-0915
Size	p. 1341-1346.
Publishing place	Elsevier Inc.
Document type	Article
ZDB-ID	189232-0
ISSN	1879-3231 ; 0093-691X
ISSN (online)	1879-3231
ISSN	0093-691X
DOI	10.1016/j.theriogenology.2016.04.076
Database	NAL-Catalogue (AGRICOLA)

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Article: N-acetyl cysteine prolonged the developmental ability of mouse two-cell embryos against oxidative stress at refrigerated temperatures

Horikoshi, Yuka / Toru Takeo / Naomi Nakagata

Cryobiology. 2016 June, v. 72, no. 3

2016

Abstract: Cold storage of two-cell embryos at refrigerated temperatures is a useful means to ship genetically engineered mice. We previously reported that M2 medium maintained the developmental ability of two-cell embryos for 48 h at 4 °C, and offspring were ... ...

Abstract	Cold storage of two-cell embryos at refrigerated temperatures is a useful means to ship genetically engineered mice. We previously reported that M2 medium maintained the developmental ability of two-cell embryos for 48 h at 4 °C, and offspring were obtained from embryos transported by a courier service under refrigerated temperatures. The limitation of 48 h practically restricts the shipping destination of the embryos. To enhance the applicability of the cold-storage technique, prolonging the time to maintain developmental ability of the embryos is required. Oxidative stress may be a cause of the declining developmental ability of cold-stored embryos. However, the effect of oxidative stress on developmental ability of embryos has not been investigated. We examined intracellular glutathione (GSH) levels of cold-stored two-cell embryos to evaluate the effect of oxidative and investigated the efficacy of adding N-acetyl cysteine (NAC) to the preservation medium on the developmental ability of cold-stored embryos and transported two-cell embryos at refrigerated temperatures. Intracellular GSH levels of two-cell embryos decreased by cold storage for longer than 72 h, whereas NAC recovered this reduction and improved the developmental ability of embryos cold-stored for 96 h. In the transport experiment, the developmental rate of transported two-cell embryos to offspring was increased by adding NAC to the preservation medium. We found that NAC prolonged the storage period of two-cell embryos and maintained the developmental ability by alleviating the reduction of intracellular GSH. These findings will improve the technique of cold-storage of two-cell embryos to facilitate efficient transport of genetically engineered mice worldwide.
Keywords	cold storage ; cysteine ; genetic engineering ; glutathione ; mice ; oxidation ; oxidative stress ; progeny ; storage time ; temperature
Language	English
Dates of publication	2016-06
Size	p. 198-204.
Publishing place	Elsevier Inc.
Document type	Article
ZDB-ID	80098-3
ISSN	1090-2392 ; 0011-2240
ISSN (online)	1090-2392
ISSN	0011-2240
DOI	10.1016/j.cryobiol.2016.05.002
Database	NAL-Catalogue (AGRICOLA)

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Article ; Online: An acute phase protein α1-acid glycoprotein mitigates AKI and its progression to CKD through its anti-inflammatory action

Hiroshi Watanabe / Rui Fujimura / Yuto Hiramoto / Ryota Murata / Kento Nishida / Jing Bi / Tadashi Imafuku / Hisakazu Komori / Hitoshi Maeda / Ayumi Mukunoki / Toru Takeo / Naomi Nakagata / Motoko Tanaka / Kazutaka Matsushita / Masafumi Fukagawa / Toru Maruyama

Scientific Reports, Vol 11, Iss 1, Pp 1-

2021 Volume 11

Abstract: Abstract The molecular mechanism for acute kidney injury (AKI) and its progression to chronic kidney disease (CKD) continues to be unclear. In this study, we investigated the pathophysiological role of the acute phase protein α1-acid glycoprotein (AGP) ... ...

Abstract	Abstract The molecular mechanism for acute kidney injury (AKI) and its progression to chronic kidney disease (CKD) continues to be unclear. In this study, we investigated the pathophysiological role of the acute phase protein α1-acid glycoprotein (AGP) in AKI and its progression to CKD using AGP KO mice. Plasma AGP levels in WT mice were increased by about 3.5-fold on day 1–2 after renal ischemia–reperfusion (IR), and these values then gradually decreased to the level before renal IR on day 7–14. On day 1 after renal IR, the AGP KO showed higher renal dysfunction, tubular injury and renal inflammation as compared with WT. On day 14, renal function, tubular injury and renal inflammation in WT had recovered, but the recovery was delayed, and renal fibrosis continued to progress in AGP KO. These results obtained from AGP KO were rescued by the administration of human-derived AGP (hAGP) simultaneously with renal IR. In vitro experiments using RAW264.7 cells showed hAGP treatment suppressed the LPS-induced macrophage inflammatory response. These data suggest that endogenously induced AGP in early renal IR functions as a renoprotective molecule via its anti-inflammatory action. Thus, AGP represents a potential target molecule for therapeutic development in AKI and its progression CKD.
Keywords	Medicine ; R ; Science ; Q
Subject code	616
Language	English
Publishing date	2021-04-01T00:00:00Z
Publisher	Nature Portfolio
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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