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Kenshiro Hara

Section

Graduate School of Agricultural Science

Job title

Associate Professor

Degree

博士（獣医学）（東京大学）
修士（農学）（東北大学）

researchmap

https://researchmap.jp/haraken

J-GLOBAL ID

201401062066559819

e-Rad No.

60551546

Research History 3

2015/04 - Present

Tohoku University　Graduate School of Agricultural Science
2010/08 - 2015/03

National Institute for Basic Biology
2009/04 - 2010/07

National Institute for Basic Biology

Education 3

国立大学法人東京大学　大学院農学生命科学研究科　獣医学専攻

2005/04 - 2009/03
国立大学法人東北大学　大学院農学研究科　応用生命科学専攻

2003/04 - 2005/03
国立大学法人東北大学　農学部　応用動物科学科

1999/04 - 2003/03

Committee Memberships 6

日本繁殖生物学会　プログラム委員

2019/04 - Present
日本繁殖生物学会　将来計画検討委員

2018/04 - Present
山形県研究評価委員

2017/10 - 2024/03
日本畜産学会　若手奨励・男女委員会委員

2019/04 - 2020/03
日本畜産学会　若手企画委員長

2019/04 - 2020/03
日本畜産学会　若手企画委員

2017/04 - 2020/03

Show all ︎Show first 5

Professional Memberships 2

日本繁殖生物学会
日本畜産学会

Research Interests 7

精子成熟
細胞系譜追跡
ライブイメージング
細胞移動
精子形成
精子幹細胞
精巣・精子

Research Areas 2

Life sciences / Veterinary medicine /
Life sciences / Animal production science /

Papers 55

Age‐Dependent Clonal Expansion of Non–Sperm‐Forming Spermatogonial Stem Cells in Mouse Testes

Terumichi Kawahara, Shinnosuke Suzuki, Toshinori Nakagawa, Yuki Kamo, Miki Kanouchi, Miyako Fujita, Maki Hattori, Atsuko Suzuki, Kentaro Tanemura, Shosei Yoshida, Kenshiro Hara

Aging Cell　2025/02/22
Publisher: Wiley
DOI： 10.1111/acel.70019 　

ISSN： 1474-9718

eISSN： 1474-9726

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ABSTRACT In male mammals, spermatogonial stem cells (SSCs) are essential for sustaining lifelong spermatogenesis within the testicular open niche, a unique environment that allows SSC migration over an extended niche area. As SSCs undergo continuous mitotic division, mutations accumulate and are transmitted to the descendant SSC clones. Therefore, SSC clonal fate behaviors, in terms of their efficiencies in completing spermatogenesis and undergoing expansion within the niche, influence sperm genomic diversity. We aimed to elucidate the effects of physiological aging on SSC clonal fate behavior within the testicular open niche. We used single‐cell RNA sequencing, lineage tracing, and intravital live imaging to investigate SSC behavior in aged mouse testes, where spermatogenesis, although reduced, persists. We found that undifferentiated spermatogonia maintained gene expression heterogeneity during aging. Among these, GFRα1⁺ cells, which exhibited state heterogeneity, showed accelerated proliferation and persistent motility, continuing to function as SSCs in older mice. In contrast, a subset of SSCs characterized by low Egr4 and Cops5 expression did not contribute to spermatid formation. These non–sperm‐forming SSC clones increased in proportion among the total SSC clones and expanded spatially within the testicular open niche in old mice, a phenomenon not observed in young mice. The expansion of non–sperm‐forming SSC clones in aged testes suggests that they occupy a niche space, limiting the availability of functional SSCs and potentially reducing sperm production and genetic diversity. These findings highlight age‐specific clonal characteristics as hallmarks of stem cell aging within the testicular open niche and provide novel insights into the mechanisms governing reproductive aging.
Male mice are susceptible to brain dysfunction induced by early-life acephate exposure

Takahiro Sasaki, Jahidul Islam, Kenshiro Hara, Tomonori Nochi, Kentaro Tanemura

Frontiers in Neuroscience　18　2024/07/10
Publisher: Frontiers Media SA
DOI： 10.3389/fnins.2024.1404009 　

eISSN： 1662-453X

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Background Acephate is a widely used organophosphate insecticide. Exposure to endocrine-disrupting chemicals, such as acephate, can interfere with neurodevelopment in childhood, increasing the risk of higher brain dysfunction later in life. Furthermore, brain dysfunction may be related to chemical exposure-related disturbances in the gut microbiota. However, the effects of early acephate exposure on the brains of adult males and females as well as on the adult gut environment remain poorly understood. Methods This study investigated the effects of perinatal acephate exposure on the central nervous system and gut microbiota of mice, including sex differences and environmentally relevant concentrations. C57BL/6 N pups were exposed to acephate (0, 0.3, 10, and 300 ppm) via the dam in their drinking water from embryonic day (E) 11.5 to postnatal day 14. We examined its effects on the central nervous system of adult males and females. Results In the male treatment group, impairments in learning and memory were detected. Immunohistochemical analysis revealed a decrease in SOX2-, NeuN-, DCX-, and GFAP-positive cells in the hippocampal dentate gyrus in males compared to the control group, whereas GFAP-positive cells were fewer in females. In addition, gut microbiota diversity was reduced in both sexes in the experimental group. Conclusion Our study demonstrates that the effects of early-life exposure to acephate are more pronounced in males than in females and can lead to a lasting impact on adult behavior, even at low doses, and that the gut microbiota may reflect the brain environment.
<i>Ccdc152</i> is not necessary for male fertility, but contributes to maintaining sperm morphology

Ryua HARIMA, Takahiro SASAKI, Takayuki KANEKO, Fuka ASO, Hayato TAKASHIMA, Takashi TOYAMA, Kenshiro HARA, Kentaro TANEMURA, Yoshiro SAITO

Journal of Reproduction and Development　2024
Publisher: Japanese Society of Animal Reproduction
DOI： 10.1262/jrd.2024-058 　

ISSN： 0916-8818

eISSN： 1348-4400
A single dose of clothianidin exposure induces varying sex-specific behavioral changes in adulthood depending on the developmental stage of its administration.

Kenshi Kaku, Takahiro Sasaki, Kenshiro Hara, Kentaro Tanemura

The Journal of toxicological sciences　49　(7)　301-311　2024

DOI： 10.2131/jts.49.301 　

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Clothianidin (CLO), a neonicotinoid that is widely used in forests and agricultural areas, was recently reported to cause toxicity in mammals. Although sensitivity to chemicals varies between sexes and developmental stages, studies that comprehensively evaluate both males and females are limited. Therefore, in this study we utilized murine models to compare the sex-specific differences in behavioral effects following CLO exposure at different developmental stages. We orally administered CLO to male and female mice as a single high-dose solution (80 mg/kg) during the postnatal period (2-week-old), adolescence (6-week-old), or maturity (10-week-old), and subsequently evaluated higher brain function. The behavioral battery test consisted of open field, light/dark transition, and contextual/cued fear conditioning tests conducted at three and seven months of age. After the behavioral test, the brains were dissected and prepared for immunohistochemical staining. We observed behavioral abnormalities in anxiety, spatial memory, and cued memory only in female mice. Moreover, the immunohistochemical analysis showed a reduction in astrocytes within the hippocampus of female mice with behavioral abnormalities. The behavioral abnormalities observed in female CLO-treated mice were consistent with the typical behavioral abnormalities associated with hippocampal astrocyte dysfunction. It is therefore possible that the CLO-induced behavioral abnormalities are at least in part related to a reduction in astrocyte numbers. The results of this study highlight the differences in behavioral effects following CLO exposure between sexes and developmental stages.
Early-life exposure to acephate inhibits sexual development and induces testicular and ovarian toxicity in mice

Takahiro Sasaki, Kenshiro Hara, Kentaro Tanemura

Reproductive Toxicology　121　108472-108472　2023/09
Publisher: Elsevier BV
DOI： 10.1016/j.reprotox.2023.108472 　

ISSN： 0890-6238
Testicular histone hyperacetylation in mice by valproic acid administration affects the next generation by changes in sperm DNA methylation. International-journal

Kazuya Sakai, Kenshiro Hara, Kentaro Tanemura

PloS one　18　(3)　e0282898　2023

DOI： 10.1371/journal.pone.0282898 　

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Various studies have described epigenetic inheritance through sperms. However, the detailed mechanisms remain unclear. In this study, we focused on DNA methylation in mice treated with valproic acid (VPA), an inducer of epigenomic changes, and analyzed the treatment effects on the sperm from the next generation of mice. The administration of 200 mg/kg/day VPA to mice for 4 weeks caused transient histone hyperacetylation in the testes and DNA methylation changes in the sperm, including promoter CpGs of genes related to brain function. Oocytes fertilized with VPA-treated mouse sperm showed methylation fluctuations at the morula stage. Pups that were fathered by these mice also showed behavioral changes in the light/dark transition test after maturation. Brain RNA-seq of these mice showed that the expression of genes related to neural functions were altered. Comparison of the sperm DNA methylation status of the next generation of mice with that of the parental generation revealed the disappearance of methylation changes observed in the sperm of the parental generation. These findings suggest that VPA-induced histone hyperacetylation may have brain function-related effects on the next generation through changes in sperm DNA methylation.
Effects of early-life tosufloxacin tosilate hydrate administration on growth rate, neurobehavior, and gut microbiota at adulthood in male mice

Ayano Hasegawa, Takahiro Sasaki, Jahidul Islam, Takashi Tominaga, Tomonori Nochi, Kenshiro Hara, Kentaro Tanemura

The Journal of Toxicological Sciences　48　(3)　149-159　2023
Publisher: Japanese Society of Toxicology
DOI： 10.2131/jts.48.149 　

ISSN： 0388-1350

eISSN： 1880-3989
Characteristics of alpha smooth muscle actin-positive peritubular cells in prepubertal bovine testes. International-journal

Yusuke Kawabe, Takashi Numabe, Kentaro Tanemura, Kenshiro Hara

Biochemical and biophysical research communications　609　48-53　2022/04/07

DOI： 10.1016/j.bbrc.2022.03.149 　

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Characteristics of peritubular myoid cells (PMCs) in sexually immature cattle remain largely unknown. Here, we report the character and behavior of peritubular cells expressing alpha smooth muscle actin (αSMA), a marker of PMCs in adult testes, in prepubertal testes procured from 5-months-old bulls. Elastin distribution around αSMA + PMCs was dim and discontinuous in prepubertal testes, but strong and continuous in adult testes. Fibroblast-specific protein 1 (FSP1) was rarely expressed in αSMA + PMCs of prepubertal testes, while in adult testes, majority of αSMA + PMCs were FSP1+. Moreover, αSMA + PMCs in prepubertal testes proliferated more actively than those in adult testes. In vitro culture of isolated seminiferous tubules from prepubertal testes revealed that αSMA + PMCs migrate from peritubular to interstitial area. Hence, in prepubertal bovine testes, (1) PMCs rarely portray fibroblast-like properties, (2) PMCs exhibit heterogeneity in FSP1 expression, (3) PMCs proliferate more actively than those in adult testes, and (4) PMCs have a potential to migrate to the interstitium. Our observations help to understand the maturation of PMCs and their involvement in bovine testicular function.
High concentration of dopamine treatment may induce acceleration of human sperm motility.

Hiroki Kanno, Shouhei Kurata, Yuuki Hiradate, Kenshiro Hara, Hiroaki Yoshida, Kentaro Tanemura

Reproductive medicine and biology　21　(1)　e12482　2022

DOI： 10.1002/rmb2.12482 　

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PURPOSE: In humans, catecholamines (including dopamine) have been identified in semen and fallopian tubes, while dopamine D2 receptors (D2DR) are found in the sperm midpiece region. How dopamine dose affects human sperm function and whether dopamine treatment is useful in assisted reproductive technology is unclear. METHODS: Sperm samples were obtained from patients with normal semen parameters undergoing fertility treatment. We investigated the effects of dopamine treatment on tyrosine phosphorylation and sperm motility. Sperm motility was analyzed using the computer-assisted sperm analysis (CASA) system. RESULTS: This study revealed that various dopamine concentrations (0.1-100 μM) did not increase sperm tyrosine phosphorylation. Progressive motility increased substantially when treated with high concentrations of dopamine (10 and 100 μM) and was blocked by raclopride (a D2DR antagonist). After 24-h sperm culture, the addition of 10 μM dopamine significantly increased curvilinear velocity and amplitude of lateral head displacement, which are indicators of hyperactivation. CONCLUSION: Dopamine did not affect tyrosine phosphorylation, but increased sperm motility. High concentrations of dopamine were more effective to accelerate sperm motility in cases where sperm motile capacity was low.
Exogenous gamma-aminobutyric acid addition enhances porcine sperm acrosome reaction. International-journal

Shouhei Kurata, Kohei Umezu, Hironori Takamori, Yuuki Hiradate, Kenshiro Hara, Kentaro Tanemura

Animal science journal = Nihon chikusan Gakkaiho　93　(1)　e13744　2022/01

DOI： 10.1111/asj.13744 　

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The widely used porcine artificial insemination procedure involves the use of liquid-stored semen because it is difficult to control the quality of frozen-thawed porcine sperm. Therefore, there is a high demand for porcine semen. The control and enhancement of sperm function are required for the efficient reproduction of pigs. We previously reported that gamma-aminobutyric acid (GABA) enhanced sperm capacitation and acrosome reaction in mice. In this study, we demonstrated the presence of GABAA receptors in porcine sperm acrosome. Furthermore, we investigated the GABA effects on porcine sperm function. We did not detect any marked effect of GABA on sperm motility and tyrosine phosphorylation of sperm proteins. However, GABA promoted acrosome reaction, which was suppressed by a selective GABAA receptor antagonist. GABA binds to GABAA receptors, resulting in chloride ion influx. We found that treatment with 1 μM GABA increased the intracellular concentration of chloride ion in the sperm. In addition, the GABA concentration effective in the acrosome reaction was correlated with the porcine sperm concentration. These results indicate that GABA and its receptors can act as modulators of acrosome reaction. This study is the first to report the effects of GABA on porcine sperm function.
Loss of Axdnd1 causes sterility due to impaired spermatid differentiation in mice

Yuki Hiradate, Ryua Harima, Rin Yanai, Kenshiro Hara, Kazue Nagasawa, Makoto Osada, Tomoe Kobayashi, Makoto Matsuyama, Shin‐ichiro Kanno, Akira Yasui, Kentaro Tanemura

Reproductive Medicine and Biology　21　(1)　2022/01
Publisher: Wiley
DOI： 10.1002/rmb2.12452 　

ISSN： 1445-5781

eISSN： 1447-0578
Potential of sperm small non-coding RNAs as biomarkers of testicular toxicity in a doxorubicin-induced mouse model. International-journal

Kazuya Sakai, Yuki Hiradate, Kenshiro Hara, Kentaro Tanemura

Biochemistry and biophysics reports　28　101160-101160　2021/12

DOI： 10.1016/j.bbrep.2021.101160 　

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Testicular toxicity is a major concern in cancer chemotherapy and drug development as it can result in infertility; however, there are no effective biomarkers for this adverse effect. To identify new biomarkers, we investigated the expression of small non-coding RNAs (sncRNAs) in a mouse model of doxorubicin (DXR)-induced testicular toxicity. First, we performed small RNA-seq analysis of sperm from DXR-treated or control mice and observed differential expression of many genome-derived sequences. We then performed real-time RT-PCR validation of these sequences and discovered that sncRNA detected by one primers, dxRN_3, showed similar differential expression as that seen in the RNA-seq experiment. These findings suggest that the sncRNAs present in sperm have potential as clinically acceptable biomarkers for testicular toxicity.
Persistence of undifferentiated spermatogonia in aged Japanese Black cattle. International-journal

Terumichi Kawahara, Miki Kanouchi, Yousuke Naniwa, Masanori Koyago, Takashi Numabe, Keishi Mizutani, Kentaro Tanemura, Kenshiro Hara

Animal science journal = Nihon chikusan Gakkaiho　92　(1)　e13572　2021/12

DOI： 10.1111/asj.13572 　

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Aging is a major risk factor for spermatogenesis deterioration. However, the influence of age on spermatogenic stem cells and their progenitors in bulls is largely unknown. Here, we report age-related changes in undifferentiated and differentiating spermatogonia in Japanese Black cattle with nearly constant sperm output, by using spermatogonial markers. The numbers of differentiating spermatogonia and more differentiated spermatogenic cells were significantly decreased in aged bovine testes compared with those in young testes. In contrast, the number of undifferentiated spermatogonia was maintained, and their proliferative activity did not differ significantly between young and aged bovine testes. Although severe calcification was only observed to a small extent in aged testes, fewer Sertoli cells and interstitial fibrosis were observed in noncalcified testicular regions. These results suggest that, even in old bulls with nearly constant sperm output, testicular spermatogenic activity declined whereas undifferentiated spermatogonia numbers were maintained. Thus, we propose that undifferentiated spermatogonia may be resistant to age-related changes in bovine testes. Because undifferentiated spermatogonia may contain stem cell activity, our findings highlight the potential utility of undifferentiated spermatogonia as an agricultural resource to produce spermatozoa beyond the natural bovine lifetime through transplantation and in vitro spermatogenesis in future animal production.
Caffeine induces sperm detachment from sperm head-to-head agglutination in bull. International-journal

Kohei Umezu, Shouhei Kurata, Kenshiro Hara, Kentaro Tanemura

Biochemical and biophysical research communications　562　105-111　2021/07/12

DOI： 10.1016/j.bbrc.2021.05.060 　

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Sperm head-to-head agglutination is a well-known known phenomenon in mammalian and non-mammalian species. Although several factors have been reported to induce sperm agglutination, information on the trigger and process of sperm detachment from the agglutination is scarce. Since hyperactivated motility is involved in bovine sperm detachment from the oviduct, we focused on caffeine, a well-known hyperactivation inducer, and aimed to determine the role of caffeine in sperm detachment from agglutination. Agglutination rate of bovine sperm was significantly decreased upon incubation with caffeine following pre-incubation without caffeine. Additionally, we observed that bovine sperm were detached from agglutination only when the medium contained caffeine. The detached sperm showed more asymmetrical flagellar beating compared to the undetached motile sperm, regardless of whether before or after the detachment. Intriguingly, some sperm that detached from agglutination re-agglutinated with different sperm agglutination. These findings indicated caffeine as a trigger for sperm detachment from the agglutination in bull. Furthermore, another well-known hyperactivation inducer, thimerosal, also significantly reduced the sperm agglutination rate. Overall, the study demonstrated the complete process of sperm detachment from sperm head-to-head agglutination and proposed that hyperactivated motility facilitates sperm detachment from another sperm. These findings would provide a better understanding of sperm physiology and fertilization process in mammals.
Behavioural effects in mice orally exposed to domoic acid or ibotenic acid are influenced by developmental stages and sex differences

Takahiro Sasaki, Hirokatsu Saito, Yuuki Hiradate, Kenshiro Hara, Kentaro Tanemura

Biochemical and Biophysical Research Communications　558　175-182　2021/06
Publisher: Elsevier BV
DOI： 10.1016/j.bbrc.2021.04.080 　

ISSN： 0006-291X
Expression and localization of alpha-tubulin N-acetyltransferase 1 in the reproductive system of male mice.

Rin Yanai, Yudai Yamashita, Kohei Umezu, Yuuki Hiradate, Kenshiro Hara, Kentaro Tanemura

The Journal of reproduction and development　67　(1)　59-66　2021/02/15

DOI： 10.1262/jrd.2020-110 　

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The structure of microtubules is essential for the fertilizing ability of spermatozoa. Acetylation of α-tubulin plays an important role in flagellar elongation and spermatozoa motility. Previous reports have suggested that alpha-tubulin N-acetyltransferase 1 (ATAT1) is the main acetyltransferase involved in the acetylation of α-tubulin. Although ATAT1 is reported to express in the testis, no information is available regarding its expression in elongated spermatids, epididymis, and mature spermatozoa. Hence, it remains unclear whether ATAT1 is involved in spermatozoa maturation and capacitation. Therefore, we evaluated the expression of ATAT1 in the mouse male reproductive system using immunostaining and western blotting. Our results showed that ATAT1 was expressed in spermatids during spermiogenesis in mouse testes, but its expression varied according to the seminiferous tubule stage. We observed ATAT1 in the cytoplasm of round spermatids, the flagella of elongated spermatids, and in the cytoplasm of step 16 spermatids, just before its release into the lumen. In addition, ATAT1 was expressed in epithelial cells of the epididymis. In spermatozoa of the cauda epididymis, ATAT1 expression was primarily observed in the midpiece of the spermatozoa. The localization of ATAT1 protein in the male germline was observed during spermiogenesis as well as during spermatozoa maturation. Our results suggest that ATAT1 may be involved in the formation of flagella and in the acetylation process, which has attracted attention in recent years regarding male infertility.
Expression and Possible Role of Nicotinic Acetylcholine Receptor ε Subunit (AChRe) in Mouse Sperm. International-journal

Yusei Makino, Yuuki Hiradate, Kohei Umezu, Kenshiro Hara, Kentaro Tanemura

Biology　10　(1)　2021/01/11

DOI： 10.3390/biology10010046 　

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The nicotinic acetylcholine receptor (nAChR) is one of the receptors of acetylcholine (ACh), and nicotine (NIC) acts as an agonist of this receptor. Among the nAChR subunits, we found that the ε subunit (AChRe) had approximately 10 to 1000 times higher level of mRNA expression in mouse testes than the other subunits. In this study, we aimed to elucidate the expression and localization of AChRe in the testes and spermatozoa of mice and clarify the effect of AChRe on sperm function. Immunocytochemistry showed that AChRe was expressed in the murine testes and spermatozoa. We found that AChRe was localized only in elongated spermatids from step 12 onwards in the testes. In spermatozoa, AChRe was localized in the head, especially in the anterior region of the acrosome, but only approximately 50% of spermatozoa showed this immunoreactivity. Additionally, we analyzed the effects of ACh and NIC on sperm acrosome reaction (AR) and found that both ACh and NIC suppressed the AR rate, which was restored by an AChRe-specific antagonist. These results suggest that AChRe may be a regulator of mammalian sperm AR.
Early life exposure to acephate exerts the neurobehavior and gut microbiota in adult mice

SASAKI Takahiro, HASEGAWA Ayano, SAKAI Kazuya, HIRADATE Yuuki, HARA Kenshiro, ISLAM Jahidul, NOCHI Tomonori, TANEMURA Kentaro

Annual Meeting of the Japanese Society of Toxicology　48　P-107S　2021
Publisher: The Japanese Society of Toxicology
DOI： 10.14869/toxpt.48.1.0_P-107S 　

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<p>Acephate (Ace), an organophosphate insecticide, is also toxic to mammals. In this study, we examined the effects of chronic Ace exposure in early life on the brain functions and gut microbiota in adult male and female mice, including acceptable daily intake (ADI) level concentrations. These results suggested that chronic exposure to Ace during the early developmental stages, even at ADI level concentrations, leads to neurobehavioral diseases and disruption of the gut microbiota. The correlation between Ace-induced behavioral effects and an altered microbiome will be further investigated.</p>
Effect of vitamin E deficiency on spermatogenesis in mice and its similarity to aging. International-journal

Hirokatsu Saito, Kenshiro Hara, Satoshi Kitajima, Kentaro Tanemura

Reproductive toxicology (Elmsford, N.Y.)　98　225-232　2020/12

DOI： 10.1016/j.reprotox.2020.10.003 　

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Vitamin E (VE) plays numerous important roles in mammals because of its antioxidant activity. As a result, VE deficiency (VED) leads to the dysfunction of central nervous, reproductive, and immune systems. However, few studies have reported the effects of VED on the male reproductive system. In this study, we investigated the effects of VED on male reproductive function and examined its relationship to involution in the male reproductive system with aging. We fed a VED or control diet to 4-week-old mice for 12 or 24 weeks. Following the histopathological analysis of reproductive organs, we found seminiferous tubules with exfoliation in the VED groups, and its frequency was significantly increased compared with the controls. Additionally, in the epididymis, a decrease in spermatozoa and an increase in apoptotic germ cells were observed in the VED groups compared with the controls. By Papanicolaou staining, we also found an increase in the proportion of sperm with abnormal morphology in the VED groups compared with the controls. These reproductive effects induced by VED were highly similar to one aspect of those observed in aged mice. Our findings demonstrate that the aging of the male reproductive system may be accelerated because of the impaired in vivo antioxidant capacity induced by VED.
Effect of neurotensin on cultured mouse preimplantation embryos. Peer-reviewed

Yuki Hiradate, Kenshiro Hara, Kentaro Tanemura

The Journal of reproduction and development　66　(5)　421-425　2020/10/13

DOI： 10.1262/jrd.2020-002 　

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Previously, we revealed that neurotensin (NTS) derived from the oviduct and uterus can function during fertilization. However, little is known about NTS actions on the pre-implantation embryo after fertilization. Here, we found that pro-Nts mRNA is expressed in the oviduct and uterus during when preimplantation embryos develop and an increase in mRNA level in the uterus is induced by human chorionic gonadotropin (hCG) treatment. Expression of mRNA for two NTS receptors, Ntr1 and Ntr3, was found throughout these stages, whereas Ntr2 mRNA was not detected, suggesting that NTS signaling occurred through NTR1 and NTR3. Supplementation of 1, 10, 100 or 1000 nM NTS to embryo culture medium after fertilization showed that 100 nM NTS significantly improved the blastocyst formation. In comparison, the total number of cells and inner cell mass ratio of blastocysts was not significant different between the 0 nM and 100 nM NTS treatment groups. These results indicate that NTS has a positive effect upon preimplantation embryo development in vitro.
Characteristics and Possible Role of Bovine Sperm Head-to-Head Agglutination. International-journal Peer-reviewed

Kohei Umezu, Shouhei Kurata, Hironori Takamori, Takashi Numabe, Yuuki Hiradate, Kenshiro Hara, Kentaro Tanemura

Cells　9　(8)　2020/08/09

DOI： 10.3390/cells9081865 　

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Although sperm head-to-head agglutination has been reported in many mammalian species, the biological significance of this unique sperm-sperm interaction remains largely unknown. Here, we aimed to examine the functional characteristics of agglutinated bovine sperm to determine the possible role of sperm agglutination in the fertilization process. We initially examined temporal changes to the degree of head-to-head agglutination in culture, and found that bovine sperm agglutinated despite the lack of sperm agglutination inducers in medium. Sperm viability and motility were evaluated by SYBR14/PI and JC-1 staining, respectively, to identify the relationship between sperm agglutination and fertilizing ability. Agglutinated sperm had increased motility, viability, and intact mitochondrial function compared with unagglutinated sperm. Furthermore, we found that heparin significantly increased the percentage of unagglutinated sperm, but did not affect viability of both agglutinated and unagglutinated sperm, suggesting that sperm agglutination dictated the viability. In conclusion, agglutinated bovine sperm maintained viability and motility for a longer time than unagglutinated sperm. Thus, we propose that the head-to-head agglutination is a crucial sperm-sperm interaction to ensure the fertilizing ability of sperm.
Spermatogonial asynchrony in Tex14 mutant mice lacking intercellular bridges. International-journal

C A Rezende-Melo, A L Caldeira-Brant, A L Drumond-Bock, G M Buchold, G Shetty, F R C L Almeida, M M Matzuk, K Hara, S Yoshida, M L Meistrich, H Chiarini-Garcia

Reproduction (Cambridge, England)　160　(2)　205-215　2020/08

DOI： 10.1530/REP-20-0118 　

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The existence of cytoplasmic passages between germ cells and their potential function in the control of the spermatogenic process has long been an intriguing question. Evidence of the important role of such structures, known as intercellular bridges (ICB), in spermatogenesis has been implicated by the failure of spermatogenesis in testis-expressed gene 14 (Tex14) mutant mice, which lack the ICBs, to progress past the pachytene spermatocyte stage. Using these Tex14 mutants, the present study evaluated, for the first time, the behavior and synchrony of the spermatogonial lineage in the absence of ICBs. Our data suggest that the absence of these cytoplasmic connections between cells affects the expansion of the undifferentiated type A (Aundiff) spermatogonia compartment and their transition to A1, resulting in a significant numerical reduction of differentiating A1 spermatogonia, but did not interfere with cell amplification during subsequent mitotic steps of differentiating spermatogonia from A1 through intermediate (In). However, beginning at the type B spermatogonia, the synchrony of differentiation was impaired as some cells showed delayed differentiation compared to their behavior in a normal seminiferous epithelium cycle. Thus although spermatogonial development is able to proceed, in the absence of ICBs in Tex14-/- mutants, the yield of cells, specific steps of differentiation, the synchrony of the cell kinetics, and the subsequent progression in meiosis are quantitatively lower than normal.
低用量ペルメトリンの早期慢性ばく露による成熟後の雄マウス行動影響

齊藤洋克, 原健士朗, 冨永貴志, 中島欽一, 北嶋聡, 菅野純, 種村健太郎

The Journal of Toxicological Sciences　45　(Suppl.)　S108-S108　2020/06
Publisher: (一社)日本毒性学会
ISSN： 0388-1350

eISSN： 1880-3989
Stromal cell-derived factor 1 regulates in vitro sperm migration towards the cumulus-oocyte complex in cattle. International-journal Peer-reviewed

Kohei Umezu, Kenshiro Hara, Yuuki Hiradate, Takashi Numabe, Kentaro Tanemura

PloS one　15　(4)　e0232536　2020

DOI： 10.1371/journal.pone.0232536 　

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Sperm migration towards an oocyte in the female reproductive tract is an important step for successful fertilization. Although several sperm-chemotactic factors have been identified in mammals, it is unclear whether these chemoattractants contribute to sperm migration towards an oocyte that is the final destination for sperm. Furthermore, chemoattractants for bovine sperm are still undiscovered even though the follicular fluid attracts sperm in cattle. Here, we demonstrated that a single bovine cumulus-oocyte complex (COC) had the ability to attract sperm, suggesting that the COC secreted sperm chemoattractants. We identified stromal cell-derived factor 1 (SDF1), which was expressed in COCs, and its receptor CXCR4 in sperm, as a candidate. Our results showed that bovine sperm preferentially migrated to the area with a high SDF1 concentration and occasionally showed turn movements by asymmetric flagellar bends during the migration. We also demonstrated that increasing the intracellular Ca2+ concentration via Ca2+ channels was related to SDF1-induced sperm chemotaxis. Finally, a CXCR4 inhibitor significantly suppressed the in vitro bovine sperm migration towards a COC. Taken together, we propose that SDF1 is a chemotactic factor for bovine sperm to regulate their migration towards an oocyte via the CXCR4 receptor.
Early-life exposure to low levels of permethrin exerts impairments in learning and memory with the effects on neuronal and glial population in adult male mice. International-journal Peer-reviewed

Hirokatsu Saito, Kenshiro Hara, Takashi Tominaga, Kinichi Nakashima, Kentaro Tanemura

Journal of applied toxicology : JAT　39　(12)　1651-1662　2019/12

DOI： 10.1002/jat.3882 　

ISSN： 0260-437X

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Permethrin, a pyrethroid chemical, is widely used as a pesticide because of its rapid insecticidal activity. Although permethrin is considered to exert very low toxicity in mammals, the effects of early, low-level, chronic exposure on the adult central nervous system are unclear. In this study, we investigated the effects of low-level, chronic permethrin exposure in early life on the brain functions of adult mice, using environmentally relevant concentrations. We exposed mice to the acceptable daily intake level of permethrin (0.3 ppm) in drinking water during the prenatal and postnatal periods. We then examined the effects on the central nervous system in adult male offspring. In the permethrin group, we detected behavior that displayed incomplete adaptation to a novel environment, as well as an impairment in learning and memory. In addition, immunohistochemical analysis revealed an increase in doublecortin- (an immature neuron marker) positive cells in the hippocampal dentate gyrus in the permethrin exposure group compared with the control group. Additionally, in the permethrin exposure group there was a decrease in astrocyte number in the hilus of the dentate gyrus, and remaining astrocytes were often irregularly shaped. These results suggest that exposure to permethrin at low levels in early life affects the formation of the neural circuit base and behavior after maturation. Therefore, in the central nervous system of male mice, low-level, chronic permethrin exposure during the prenatal and postnatal periods has effects that were not expected based on the known effects of permethrin exposure in mature animals.
Capacitation of mouse sperm is modulated by gamma-aminobutyric acid (GABA) concentration. Peer-reviewed

Shouhei Kurata, Yuuki Hiradate, Kohei Umezu, Kenshiro Hara, Kentaro Tanemura

The Journal of reproduction and development　65　(4)　327-334　2019/08/09

DOI： 10.1262/jrd.2019-008 　

ISSN： 0916-8818

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In mammals, ejaculated sperm acquire their fertilizing ability during migration through the female reproductive tract, which secretes several factors that contribute to sperm capacitation. Gamma-aminobutyric acid (GABA) is a well-known neurotransmitter in the central nervous system, but additionally enhances the sperm acrosome reaction in the rat and cow. However, the detailed effects of GABA concentration on sperm function remain unclear. In this study, we detected the presence of the GABA type A receptor (GABA A) in mouse epididymal sperm by western blot analysis and in the sperm acrosome by immunocytochemistry. We also investigated the effects of GABA on sperm fertilizing ability. We found that GABA facilitated the tyrosine phosphorylation of sperm proteins, which is an index of sperm capacitation. GABA also promoted the acrosome reaction, which was suppressed by a selective GABA A receptor antagonist. We then found that the effective GABA concentration for the acrosome reaction corresponds to sperm concentration, but we did not detect any marked effect of GABA on sperm motility using a computer-assisted sperm analysis system. Using immunohistochemistry, we also detected GABA expression in the epithelia of the mouse uterus and oviduct. Furthermore, we found that the mRNA levels of glutamate decarboxylase (Gad), which generates GABA from L-glutamate, were higher in the oviduct than in the uterus, and that Gad mRNA levels were higher at estrus than at the diestrus stage. These results indicate that the GABA concentration can act as a modulator of the acrosome reaction and sperm capacitation in the female reproductive tract.
Differences in resistance against osmotic challenge among C57BL/6, DBA/2 and their hybrid mice metaphase II (MII) stage oocytes. International-journal Peer-reviewed

Mei Goto, Hirokatsu Saito, Yuki Hiradate, Kenshiro Hara, Kentaro Tanemura

Zygote (Cambridge, England)　27　(4)　250-254　2019/08

DOI： 10.1017/S0967199418000370 　

ISSN： 0967-1994

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Oocytes of B6D2F1 (BDF1) mice are often used as recipients for intracytoplasmic sperm injection because of their cell membrane resistance against capillary penetration. It is assumed that oocytes of BDF1 mice have superior traits because of their hybrid vigour. However, the mechanisms of hybrid vigour are unclear. In this study, we focused on the membrane resistance of MII stage oocytes against changes in extracellular osmotic pressure. As a result, MII stage oocytes of inbred C57BL/6 and DBA/2 mice showed high tolerance in either a hypertonic or a hypotonic environment. Conversely, MII stage oocytes of hybrid BDF1 and D2B6F1 mice showed high tolerance in both hypertonic and hypotonic environments. Therefore, it is considered that MII stage oocytes of hybrid mice have superior traits than those of inbred mice. Our findings demonstrated that the hybrid vigour exists in the form of resistance to extracellular osmotic environment in hybrid MII stage oocytes.
Mammalian germ cell migration during development, growth, and homeostasis. Peer-reviewed

Mizuho Kanamori, Kenta Oikawa, Kentaro Tanemura, Kenshiro Hara

Reproductive medicine and biology　18　(3)　247-255　2019/07

DOI： 10.1002/rmb2.12283 　

ISSN： 1445-5781

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Background: Germ cells represent one of the typical cell types that moves over a long period of time and large distance within the animal body. To continue its life cycle, germ cells must migrate to spatially distinct locations for proper development. Defects in such migration processes can result in infertility. Thus, for more than a century, the principles of germ cell migration have been a focus of interest in the field of reproductive biology. Methods: Based on published reports (mainly from rodents), investigations of germ cell migration before releasing from the body, including primordial germ cells (PGCs), gonocytes, spermatogonia, and immature spermatozoon, were summarized. Main findings: Germ cells migrate with various patterns, with each migration step regulated by distinct mechanisms. During development, PGCs actively and passively migrate from the extraembryonic region toward genital ridges through the hindgut epithelium. After sex determination, male germline cells migrate heterogeneously in a developmental stage-dependent manner within the testis. Conclusion: During migration, there are multiple gates that disallow germ cells from re-entering the proper developmental pathway after wandering off the original migration path. The presence of gates may ensure the robustness of germ cell development during development, growth, and homeostasis.
Improvement in blastocyst quality by neurotensin signaling via its receptors in bovine spermatozoa during in vitro fertilization. Peer-reviewed

Kohei Umezu, Risa Yajima, Yuuki Hiradate, Rin Yanai, Takashi Numabe, Kenshiro Hara, Toshinori Oikawa, Kentaro Tanemura

The Journal of reproduction and development　65　(2)　147-153　2019/04/12

DOI： 10.1262/jrd.2018-147 　

ISSN： 0916-8818
Effects of doxorubicin on sperm DNA methylation in mouse models of testicular toxicity. International-journal Peer-reviewed

Kazuya Sakai, Maky Ideta-Otsuka, Hirokatsu Saito, Yuki Hiradate, Kenshiro Hara, Katsuhide Igarashi, Kentaro Tanemura

Biochemical and biophysical research communications　498　(3)　674-679　2018/04/06

DOI： 10.1016/j.bbrc.2018.03.044 　

ISSN： 0006-291X

eISSN： 1090-2104
Potassium bromate disrupts mitochondrial distribution within murine oocytes during in vitro maturation. Peer-reviewed

Kenichi Yamada, Yuuki Hiradate, Mei Goto, Chiho Nishiyama, Kenshiro Hara, Hiroaki Yoshida, Kentaro Tanemura

Reproductive medicine and biology　17　(2)　143-148　2018/04

DOI： 10.1002/rmb2.12079 　

ISSN： 1445-5781

eISSN： 1447-0578
Prolyl isomerase Pin1 is required sperm production by promoting mitosis progression of spermatogonial stem cells. International-journal Peer-reviewed

Atsuko Kurita-Suzuki, Yuki Kamo, Chiyoko Uchida, Kentaro Tanemura, Kenshiro Hara, Takafumi Uchida

Biochemical and biophysical research communications　497　(1)　388-393　2018/02/26
Publisher: Elsevier B.V.
DOI： 10.1016/j.bbrc.2018.02.090 　

ISSN： 1090-2104 0006-291X
Prenatal and postnatal exposure to low levels of permethrin exerts reproductive effects in male mice. International-journal Peer-reviewed

Hirokatsu Saito, Kenshiro Hara, Kentaro Tanemura

Reproductive toxicology (Elmsford, N.Y.)　74　108-115　2017/12

DOI： 10.1016/j.reprotox.2017.08.022 　

ISSN： 0890-6238
Effects on glycocalyx structures of frozen-thawed bovine sperm induced by flow cytometry and artificial capacitation. Peer-reviewed

Kohei Umezu, Yuuki Hiradate, Takashi Numabe, Kenshiro Hara, Kentaro Tanemura

The Journal of reproduction and development　63　(5)　473-480　2017/10/18

DOI： 10.1262/jrd.2017-065 　

ISSN： 0916-8818
SHISA6 Confers Resistance to Differentiation-Promoting Wnt/β-Catenin Signaling in Mouse Spermatogenic Stem Cells. International-journal Peer-reviewed

Moe Tokue, Kanako Ikami, Seiya Mizuno, Chiyo Takagi, Asuka Miyagi, Ritsuko Takada, Chiyo Noda, Yu Kitadate, Kenshiro Hara, Hiroko Mizuguchi, Takuya Sato, Makoto Mark Taketo, Fumihiro Sugiyama, Takehiko Ogawa, Satoru Kobayashi, Naoto Ueno, Satoru Takahashi, Shinji Takada, Shosei Yoshida

Stem cell reports　8　(3)　561-575　2017/03/14

DOI： 10.1016/j.stemcr.2017.01.006 　

ISSN： 2213-6711
Exogenous neurotensin modulates sperm function in Japanese Black cattle. Peer-reviewed

Kohei Umezu, Yuuki Hiradate, Toshinori Oikawa, Hirotoshi Ishiguro, Takashi Numabe, Kenshiro Hara, Kentaro Tanemura

The Journal of reproduction and development　62　(4)　409-14　2016/08/25

DOI： 10.1262/jrd.2016-055 　

ISSN： 0916-8818
Genome-Scale Assessment of Age-Related DNA Methylation Changes in Mouse Spermatozoa. International-journal Peer-reviewed

Norio Kobayashi, Hiroaki Okae, Hitoshi Hiura, Hatsune Chiba, Yoshiki Shirakata, Kenshiro Hara, Kentaro Tanemura, Takahiro Arima

PloS one　11　(11)　e0167127　2016

DOI： 10.1371/journal.pone.0167127 　

ISSN： 1932-6203
Hierarchical differentiation competence in response to retinoic acid ensures stem cell maintenance during mouse spermatogenesis. International-journal Peer-reviewed

Kanako Ikami, Moe Tokue, Ryo Sugimoto, Chiyo Noda, Satoru Kobayashi, Kenshiro Hara, Shosei Yoshida

Development (Cambridge, England)　142　(9)　1582-92　2015/05/01

DOI： 10.1242/dev.118695 　

ISSN： 0950-1991

eISSN： 1477-9129
Mouse Spermatogenic Stem Cells Continually Interconvert between Equipotent Singly Isolated and Syncytial States Peer-reviewed

Kenshiro Hara, Toshinori Nakagawa, Hideki Enomoto, Mikiko Suzuki, Masayuki Yamamoto, Benjamin D. Simons, Shosei Yoshida

CELL STEM CELL　14　(5)　658-672　2014/05

DOI： 10.1016/j.stem.2014.01.019 　

ISSN： 1934-5909

eISSN： 1875-9777
Sox17 haploinsufficiency results in perinatal biliary atresia and hepatitis in C57BL/6 background mice Peer-reviewed

Mami Uemura, Aisa Ozawa, Takumi Nagata, Kaoruko Kurasawa, Naoki Tsunekawa, Ikuo Nobuhisa, Tetsuya Taga, Kenshiro Hara, Akihiko Kudo, Hayato Kawakami, Yukio Saijoh, Masamichi Kurohmaru, Masami Kanai-Azuma, Yoshiakira Kanai

DEVELOPMENT　140　(3)　639-648　2013/02

DOI： 10.1242/dev.086702 　

ISSN： 0950-1991
マウス精子幹細胞の多様な細胞運命

原健士朗, 吉田松生

日本繁殖生物学会講演要旨集　106　OR2-3-OR2-3　2013
Publisher: 日本繁殖生物学会
DOI： 10.14882/jrds.106.0.OR2-3.0 　

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【背景】哺乳類の精子幹細胞は，繁殖期間を通じて「自己複製」と「分化細胞の供給」を両立し続けることによって恒常的な精子形成を支える。これがバランスを崩した場合，腫瘍化もしくは幹細胞の枯渇に至り，精子形成を維持できない。このことから，幹細胞の自己複製と分化パターンの理解は，精子形成における重要な課題のひとつといえる。しかし，これまで個々の精子幹細胞がどのような運命を辿って精子形成を支えているかは殆ど不明であった。そこで本研究では，マウスをモデルとして，精子幹細胞の辿る運命の解明を試みた。【材料と方法】3か月齢のGFRα1-CreER; CAG-CAT-GFPマウスの腹腔内にタモキシフェン0.35㎎を投与し，GFRα1発現細胞を不可逆的にGFPラベルした後，2，4，6，10，14，20日経過後に精巣をサンプリングした。単離した精細管を固定後，抗GFP抗体および抗GFRα1抗体を用いて免疫組織化学染色を施し，各クローン（GFP発現細胞細胞群）に含まれる自己複製（GFRα1発現を維持）した細胞と，分化（GFRα1発現を消失）した細胞の数を計測した。【結果と考察】個々の幹細胞クローンは，全て自己複製したもの，一部が自己複製して残りは分化したもの，または全て分化したものなど，それぞれの運命は多様であった。また，クローン運命の多様性は時間を経るごとに増大した。これに対し，各クローンのデータを細胞集団レベル（全クローンのデータの平均）でみると，GFRα1発現細胞は，自己の数を一定に保ちつつ，同時に分化細胞を生み出し続ける幹細胞集団であることを見出した。以上の結果，個々の精子幹細胞は多様な運命を辿りながら精子形成を支えていることが示唆された。本会では，精子幹細胞の運命生成の原理についても現在進行中の解析結果を交えて議論したい。
Effects of ubiquitin C-terminal hydrolase L1 deficiency on mouse ova Peer-reviewed

Sayaka Koyanagi, Hiroko Hamasaki, Satoshi Sekiguchi, Kenshiro Hara, Yoshiyuki Ishii, Shigeru Kyuwa, Yasuhiro Yoshikawa

REPRODUCTION　143　(3)　271-279　2012/03

DOI： 10.1530/REP-11-0128 　

ISSN： 1470-1626
Cyclical and Patch-Like GDNF Distribution along the Basal Surface of Sertoli Cells in Mouse and Hamster Testes Peer-reviewed

Takeshi Sato, Yoshimi Aiyama, Mayuko Ishii-Inagaki, Kenshiro Hara, Naoki Tsunekawa, Kyoko Harikae, Mami Uemura-Kamata, Mai Shinomura, Xiao Bo Zhu, Seishi Maeda, Sachi Kuwahara-Otani, Akihiko Kudo, Hayato Kawakami, Masami Kanai-Azuma, Michio Fujiwara, Yoichi Miyamae, Shosei Yoshida, Makoto Seki, Masamichi Kurohmaru, Yoshiakira Kanai

PLOS ONE　6　(12)　e28367　2011/12

DOI： 10.1371/journal.pone.0028367 　

ISSN： 1932-6203
哺乳類の性分化、発生過程、性成熟、季節繁殖、冬眠状態でのセルトリ細胞のニッチ能の動態

佐藤剛, 相山好美, 稲垣麻由子, 篠村麻衣, 張替香生子, 原健士朗, 上村麻実, 恒川直樹, 宮東昭彦, 金井正美, 川上速人, 関眞, 関二郎, 九郎丸正道, 金井克晃

日本生化学会大会・日本分子生物学会年会合同大会講演要旨集　83回・33回　2P-0813　2010/12
Publisher: (公社)日本生化学会
Expression and function of mouse Sox17 gene in the specification of gallbladder/bile-duct progenitors during early foregut morphogenesis Peer-reviewed

Mami Uemura, Kenshiro Hara, Hiroshi Shitara, Rie Ishii, Naoki Tsunekawa, Yutaroh Miura, Masamichi Kurohmaru, Choji Taya, Hiromichi Yonekawa, Masami Kanai-Azuma, Yoshiakira Kanai

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS　391　(1)　357-363　2010/01

DOI： 10.1016/j.bbrc.2009.11.063 　

ISSN： 0006-291X
Evidence for crucial role of hindgut expansion in directing proper migration of primordial germ cells in mouse early embryogenesis Peer-reviewed

Kenshiro Hara, Masami Kanai-Azuma, Mami Uemura, Hiroshi Shitara, Choji Taya, Hiromichi Yonekawa, Hayato Kawakami, Naoki Tsunekawa, Masamichi Kurohmaru, Yoshiakira Kanai

DEVELOPMENTAL BIOLOGY　330　(2)　427-439　2009/06

DOI： 10.1016/j.ydbio.2009.04.012 　

ISSN： 0012-1606
Ratio of peripheral nervous tissues in tongues, skeletal muscles and intestines in cows. Peer-reviewed

XiaoBo Zhu, Shogo Matoba, Kenshiro Hara, Yasuhisa Ano, Shin-ichi Kobayashi, Naoki Tsunekawa, Yoshiakira Kanai, Noboru Manabe, Takashi Onodera, Masamichi Kurohmaru

Okajimas folia anatomica Japonica　85　(2)　73-7　2008/08
Publisher: 2
DOI： 10.2535/ofaj.85.73 　

ISSN： 0030-154X

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It is suggested that pathogenic prion protein (PrP(sc) may accumulate in peripheral nervous tissues in addition to the specified-risk materials. To assess the risk of PrP(sc) accumulation in peripheral tissues, the present study was designed to study the ratio of peripheral nervous tissues in tongues, skeletal muscles and intestines in 2-week- and 59-month-old cows. The nervous tissues could be detected by immunohistochemistry using antibody Neurofilament 150. After immunostaining with Neurofilament 150, the photographs taken from each section were applied for Scion Image software. Then, the ratio of nervous tissues in tongues, skeletal muscles and intestines were calculated. The highest ratio of peripheral nervous tissues was observed in intestines, the second highest ratio was found in tongues, and the lowest ratio was recognized in skeletal muscles in both 2-week- and 59-month-old cows. The ratio of nervous tissues in intestines was approximately 2.5 fold and 10 fold, compared to that in skeletal muscles in 2-week- and 59-month-old cows, respectively. These results suggest that intestines and tongues may have a higher risk of PrP(sc) accumulation, compared to skeletal muscles.
Sox17 plays a substantial role in late-stage differentiation of the extraembryonic endoderm in vitro Peer-reviewed

Masafumi Shimoda, Masami Kanai-Azuma, Kenshiro Hara, Satsuki Miyazaki, Yoshiakira Kanai, Morito Monden, Jun-ichi Miyazaki

JOURNAL OF CELL SCIENCE　120　(21)　3859-3869　2007/11

DOI： 10.1242/jcs.007856 　

ISSN： 0021-9533

eISSN： 1477-9137
Redundant roles of Sox17 and Sox18 in early cardiovascular development of mouse embryos Peer-reviewed

Youhei Sakamoto, Kenshiro Hara, Masami Kanai-Azuma, Toshiyasu Matsui, Yutaroh Miura, Naoki Tsunekawa, Masamichi Kurohmaru, Yukio Saijoh, Peter Koopman, Yoshiakira Kanai

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS　360　(3)　539-544　2007/08

DOI： 10.1016/j.bbrc.2007.06.093 　

ISSN： 0006-291X
Isolation, characterization, and in vitro and in vivo differentiation of putative thecal stem cells Peer-reviewed

Arata Honda, Michiko Hirose, Kenshiro Hara, Shogo Matoba, Kimiko Inoue, Himomi Miki, Hitoshi Hiura, Mito Kanatsu-Shinohara, Yoshiakira Kanai, Tomohiro Kono, Takashi Shinohara, Atsuo Ogura

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA　104　(30)　12389-12394　2007/07

DOI： 10.1073/pnas.0703787104 　

ISSN： 0027-8424
Redundant roles of Sox17 and Sox18 in postnatal angiogenesis in mice Peer-reviewed

Toshiyasu Matsui, Masami Kanai-Azuma, Kenshiro Hara, Shogo Matoba, Ryuji Hiramatsu, Hayato Kawakami, Masamichi Kurohmaru, Peter Koopman, Yoshiakira Kanai

JOURNAL OF CELL SCIENCE　119　(17)　3513-3526　2006/09

DOI： 10.1242/jcs.03081 　

ISSN： 0021-9533
Production of live offspring from mouse germinal vesicle-stage oocytes vitrified by a modified stepwise method, SWEID Peer-reviewed

N Aono, Y Abe, K Hara, H Sasada, E Sato, H Yoshida

FERTILITY AND STERILITY　84　1078-1082　2005/10

DOI： 10.1016/j.fertnstert.2005.03.077 　

ISSN： 0015-0282
Feasibility of a nylon-mesh holder for vitrification of bovine germinal vesicle oocytes in subsequent production of viable blastocysts Peer-reviewed

Y Abe, K Hara, H Matsumoto, J Kobayashi-, H Sasada, H Ekwall, H Rodriguez-Martinez, E Sato

BIOLOGY OF REPRODUCTION　72　(6)　1416-1420　2005/06

DOI： 10.1095/biolreprod.104.037051 　

ISSN： 0006-3363
Extrusion and removal of lipid from the cytoplasm of porcine oocytes at the germinal vesicle stage: Centrifugation under hypertonic conditions influences vitrification Peer-reviewed

K Hara, Y Abe, N Kumada, N Aono, J Kobayashi, H Matsumoto, H Sasada, E Sato

CRYOBIOLOGY　50　(2)　216-222　2005/04

DOI： 10.1016/j.cryobiol.2005.01.003 　

ISSN： 0011-2240
Successful production of blastocysts following ultrarapid vitrification with step-wise equilibriation of germinal VesicleStage mouse oocytes Peer-reviewed

N Aono, T Naganuma, Y Abe, K Hara, H Sasada, E Sato, H Yoshida

JOURNAL OF REPRODUCTION AND DEVELOPMENT　49　(6)　501-506　2003/12

DOI： 10.1262/jrd.49.501 　

ISSN： 0916-8818

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Misc. 52

ビタミンE欠乏給餌によるマウス雄性生殖器および精子への影響と加齢による退行変化との類似性

齊藤洋克, 原健士朗, 北嶋聡, 種村健太郎

日本食品衛生学会学術講演会講演要旨集　116th　2020
ウシにおける凝集精子の特性と役割

梅津康平, 倉田笙平, 平舘裕希, 原健士朗, 種村健太郎

Journal of Reproduction and Development　66　(Suppl Japanese Issue)　2020

ISSN： 0916-8818
Early-life exposure to low levels of permethrin exerts impairments in learning and memory with the effects on neuronal and glial population in adult male mice

齊藤洋克, 原健士朗, 冨永貴志, 中島欽一, 北嶋聡, 菅野純, 種村健太郎

Journal of Toxicological Sciences　45　(Supplement)　2020

ISSN： 0388-1350
精子エピゲノムを用いた早期精巣毒性バイオマーカーの探索

酒井和哉, 大塚まき, 斉藤洋克, 平舘裕希, 原健士朗, 五十嵐勝秀, 種村健太郎

The Journal of Reproduction and Development　65　(Suppl.)　j59-j59　2019/09
Publisher: (公社)日本繁殖生物学会
ISSN： 0916-8818
ニコチン型アセチルコリン受容体シグナルを利用したマウス精子機能の調節

牧野優誠, 平舘裕希, 矢内凛, 斉藤洋克, 原健士朗, 種村健太郎

The Journal of Reproduction and Development　65　(Suppl.)　j105-j105　2019/09
Publisher: (公社)日本繁殖生物学会
ISSN： 0916-8818
ウシ精子走化性因子の特定と制御機構の解明

梅津康平, 平舘裕希, 原健士朗, 種村健太郎

日本胚移植学雑誌　41　(1 (CD-ROM))　2019

ISSN： 1341-2965
ウシ精子を卵母細胞に導く走化性因子の特定

梅津康平, 平舘裕希, 原健士朗, 種村健太郎

Journal of Reproduction and Development　65　(Suppl Japanese Issue)　2019

ISSN： 0916-8818
γ-アミノ酪酸(GABA)を利用した精子機能制御法の開発に資する研究

倉田笙平, 平舘裕希, 梅津康平, 原健士朗, 種村健太郎

Journal of Reproduction and Development　65　(Suppl Japanese Issue)　2019

ISSN： 0916-8818
低用量ペルメトリン早期慢性暴露によるマウス次世代雄個体行動影響

斉藤洋克, 原健士朗, 冨永貴志, 中島欽一, 種村健太郎

日本内分泌かく乱化学物質学会研究発表会要旨集　21回　58-58　2018/12
Publisher: 環境ホルモン学会(日本内分泌撹乱化学物質学会)
B6マウス精巣における精子形成の加齢変化

加茂祐樹, 叶内史記, 金森瑞歩, 種村健太郎, 原健士朗

The Journal of Reproduction and Development　64　(Suppl.)　j97-j97　2018/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
黒毛和種の精巣における精子形成の形態学的加齢変化

叶内史記, 島岡良登, 石山義浩, 影山恵理, 難波陽介, 古家後雅典, 水谷啓司, 沼邊孝, 種村健太郎, 原健士朗

The Journal of Reproduction and Development　64　(Suppl.)　j98-j98　2018/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
マウス精子機能に対するGABAの濃度影響解析

倉田笙平, 梅津康平, 平舘裕希, 原健士朗, 種村健太郎

日本畜産学会大会講演要旨集　124回　209-209　2018/03
Publisher: (公社)日本畜産学会
ISSN： 1342-4688
ウシ精子に対するSDF1の機能解析

梅津康平, 矢内凜, 平舘裕希, 原健士朗, 種村健太郎

日本畜産学会大会講演要旨集　124回　209-209　2018/03
Publisher: (公社)日本畜産学会
ISSN： 1342-4688
酸化ストレスがマウス体外成熟培養の成熟率とミトコンドリアクラスターに及ぼす影響

山田健市, 吉田仁秋, 原健士朗, 種村健太郎

日本IVF学会誌　20　(2)　61-61　2017/09
Publisher: (一社)日本IVF学会
ISSN： 1881-9028
ウシ精子におけるケモカインの受容体発現解析および機能解析

梅津康平, 原健士朗, 種村健太郎

日本獣医学会学術集会講演要旨集　160回　312-312　2017/08
Publisher: (公社)日本獣医学会
ISSN： 1347-8621
マウス成熟卵母細胞におけるタウの発現およびリン酸化様式の解析

西山智穂, 平舘裕希, 原健士朗, 種村健太郎

日本獣医学会学術集会講演要旨集　160回　313-313　2017/08
Publisher: (公社)日本獣医学会
ISSN： 1347-8621
人工授精用ウシ凍結精子における糖鎖構造解析

梅津康平, 原健士朗, 種村健太郎

日本畜産学会大会講演要旨集　122回　108-108　2017/03
Publisher: (公社)日本畜産学会
ISSN： 1342-4688
ニューロテンシンはウシ凍結精液の受精能獲得および先体反応を促進する

梅津康平, 平舘裕希, 原健士朗, 種村健太郎

The Journal of Reproduction and Development　62　(Suppl.)　j61-j61　2016/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
性成熟から繁殖期を通じたマウス精子DNAメチル化様式の加齢性変化

小林記緒, 岡江寛明, 樋浦仁, 千葉初音, 白形芳樹, 原健士朗, 有馬隆博, 種村健太郎

The Journal of Reproduction and Development　62　(Suppl.)　j62-j62　2016/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
バルプロ酸投与によるマウス精子エピゲノム様式の変化

関根雅史, 小林記緒, 白形芳樹, 岡江寛明, 樋浦仁, 平舘裕希, 原健士朗, 有馬隆博, 種村健太郎

The Journal of Reproduction and Development　62　(Suppl.)　j78-j78　2016/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
ビタミンE欠乏給餌によるマウス精巣影響と精子DNAメチローム変化

斉藤洋克, 井上弘貴, 小林記緒, 白形芳樹, 岡江寛明, 樋浦仁, 原健士朗, 有馬隆博, 種村健太郎

The Journal of Reproduction and Development　62　(Suppl.)　j122-j122　2016/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
浸透圧変動に対するMII期卵母細胞の細胞膜状態についてのマウス系統間の差違

後藤萌, 斉藤洋克, 白形芳樹, 原健士朗, 種村健太郎

The Journal of Reproduction and Development　62　(Suppl.)　j123-j123　2016/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
細胞社会の理解が照らし出す組織構築原理マウス精巣における精子幹細胞の動態

原健士朗, 種村健太郎, 吉田松生

日本生化学会大会プログラム・講演要旨集　89回　[3S08-2]　2016/09
Publisher: (公社)日本生化学会
ウシ精子機能へのニューロテンシンの影響

梅津康平, 平舘裕希, 原健士朗, 種村健太郎

日本獣医学会学術集会講演要旨集　159回　315-315　2016/08
Publisher: (公社)日本獣医学会
ISSN： 1347-8621
ブタ体外成熟卵母細胞のガラス化保存におけるラクトフェリンの効果

根岸真奈美, 石川禎将, 平賀孔, 杉村智史, 原健士朗, 種村健太郎

東北畜産学会報　66　(1)　7-15　2016/06
Publisher: 東北畜産学会
ISSN： 1341-626X
ウシ精子機能に対するニューロテンシン(NT)の影響解析

梅津康平, 平舘裕希, 原健士朗, 種村健太郎

日本畜産学会大会講演要旨　121st　147　2016/03/27

ISSN： 1342-4688
繁殖期におけるマウス精子DNAメチル化様式の加齢性変化

小林記緒, 小林記緒, 岡江寛明, 樋浦仁, 千葉初音, 白形芳樹, 原健士朗, 種村健太郎, 有馬隆博

日本分子生物学会年会プログラム・要旨集(Web)　39th　ROMBUNNO.1P‐0151 (WEB ONLY)　2016
幼若期マウスへのネオニコチノイド系農薬投与による神経行動毒性発現

種村健太郎, 古川祐介, 斉藤洋克, 白形芳樹, 原健士朗, 北嶋聡, 菅野純

環境ホルモン学会研究発表会要旨集　18回　78-78　2015/12
Publisher: 環境ホルモン学会(日本内分泌撹乱化学物質学会)
社会的生育環境要因が惹起する雄性生殖細胞系列および次世代へのエピジェネティック影響

小林記緒, 白形芳樹, 平舘裕希, 岡江寛明, 樋浦仁, 原健士朗, 有馬隆博, 種村健太郎

The Journal of Reproduction and Development　61　(Suppl.)　j76-j76　2015/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
低用量ビスフェノール類慢性暴露によるマウス雄性生殖細胞エピジェネティック修飾への影響解析

白形芳樹, 小林記緒, 平舘裕希, 岡江寛明, 樋浦仁, 原健士朗, 有馬隆博, 種村健太郎

The Journal of Reproduction and Development　61　(Suppl.)　j77-j77　2015/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
幼若期雄マウスへのペルメトリン投与による成熟期生殖機能影響解析

斉藤洋克, 植松未知, 白形芳樹, 原健士朗, 種村健太郎

The Journal of Reproduction and Development　61　(Suppl.)　j78-j78　2015/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
Hanging Drop法を用いた体外成熟単培養系におけるマウス卵母細胞へのネオニコチノイド類暴露影響解析

猪股大貢, 原健士朗, 種村健太郎

The Journal of Reproduction and Development　61　(Suppl.)　j90-j90　2015/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
精細管基底区画における分化軸に沿った精原細胞の局在変化

原健士朗, 種村健太郎, 吉田松生

The Journal of Reproduction and Development　61　(Suppl.)　j131-j131　2015/09
Publisher: (一社)日本繁殖生物学会
ISSN： 0916-8818
低用量ビスフェノールAFの慢性暴露による次世代雄個体中枢影響解析

白形芳樹, 小林記緒, 平舘裕希, 原健士朗, 種村健太郎

日本獣医学会学術集会講演要旨集　158回　280-280　2015/08
Publisher: (公社)日本獣医学会
ISSN： 1347-8621
ビタミンE欠乏給餌におけるマウス雄性生殖機能影響解明

斉藤洋克, 井上弘貴, 小林記緒, 白形芳樹, 岡江寛明, 樋浦仁, 原健士朗, 有馬隆博, 種村健太郎

日本獣医学会学術集会講演要旨集　158回　281-281　2015/08
Publisher: (公社)日本獣医学会
ISSN： 1347-8621
生体内におけるマウス精子エピゲノム改変の化学的誘導

関根雅史, 小林記緒, 白形芳樹, 岡江寛明, 樋浦仁, 平舘裕希, 原健士朗, 有馬隆博, 種村健太郎

日本獣医学会学術集会講演要旨集　158回　281-281　2015/08
Publisher: (公社)日本獣医学会
ISSN： 1347-8621
マウス精子におけるMeCP2タンパクの発現解析

小林記緒, 白形芳樹, 平舘裕希, 原健士朗, 種村健太郎

日本獣医学会学術集会講演要旨集　158回　281-281　2015/08
Publisher: (公社)日本獣医学会
ISSN： 1347-8621
ウシ精子におけるレクチン結合様式

梅津康平, 平舘裕希, 原健士朗, 種村健太郎

日本獣医学会学術集会講演要旨集　158回　281-281　2015/08
Publisher: (公社)日本獣医学会
ISSN： 1347-8621
ブタ精子におけるヒストンH4の修飾動態解析

内藤秋, 白形芳樹, 小林記緒, 斉藤隼人, 沼邊孝, 平舘裕希, 原健士朗, 種村健太郎

日本獣医学会学術集会講演要旨集　158回　282-282　2015/08
Publisher: (公社)日本獣医学会
ISSN： 1347-8621
バルプロ酸の利用によるマウス精子エピゲノム改変誘導の試み

関根雅史, 小林記緒, 白形芳樹, 岡江寛明, 樋浦仁, 平舘裕希, 原健士朗, 有馬隆博, 種村健太郎

東北畜産学会報　65　(2)　17-17　2015/08
Publisher: 東北畜産学会
ISSN： 1341-626X
フローサイトメトリー分離によるウシ精子のレクチン結合様式への影響

梅津康平, 平舘裕希, 原健士朗, 種村健太郎

東北畜産学会報　65　(2)　17-17　2015/08
Publisher: 東北畜産学会
ISSN： 1341-626X
受精能獲得に伴うブタ精子のヒストンH4修飾様式の変化

内藤秋, 白形芳樹, 小林記緒, 斉藤隼人, 沼邊孝, 平舘裕希, 原健士朗, 種村健太郎

東北畜産学会報　65　(2)　18-18　2015/08
Publisher: 東北畜産学会
ISSN： 1341-626X
Stem cell functionality in the mouse spermatogenesis

Shosei Yoshida, Toshinori Nakagawa, Kenshiro Hara, Yu Kitadate

DIFFERENTIATION　80　S11-S11　2010/11

DOI： 10.1016/j.diff.2010.09.162 　

ISSN： 0301-4681
ホールマウントin situハイブリダイゼーション(B.in situハイブリダイゼーション法)

金井正美, 川上速人, 原健士朗, 金井克晃

組織細胞化学　51-58　2008/07
生殖細胞分化過程におけるHuBの発現

渡邉陽子, 米田明日香, 原健士朗, 的場章悟, 平松竜司, 星野安信, 九郎丸正道, 恒川直樹, 金井克晃

J Reprod Dev　53　(Supplement)　J174　2007/09/25

DOI： 10.14882/jrds.100.0.20066.0 　

ISSN： 0916-8818
マウス始原生殖細胞におけるHuBの発現

渡邉陽子, 原健士朗, 米田明日香, 星野安信, 三浦雄太郎, 恒川直樹, 金井克晃, 九郎丸正道

日本獣医学会学術集会講演要旨集　143rd　148　2007/03/15

ISSN： 1347-8621
マウス生殖細胞系列におけるELAV様のRNA結合蛋白Huファミリー発現

米田明日香, 渡邉陽子, 的場章悟, 原健士朗, 平松竜司, 星野安信, 九郎丸正道, 恒川直樹, 金井克晃

生化学　2P-1304　2007

ISSN： 0037-1017
The maturational competence of porcine reconstructed oocytes with germinal vesicle donors vitrified

ABE Yasuyuki, KUMADA Nobuyuki, HARA Kenshiro, MATSUMOTO Hiromichi, SASADA Hiroshi, SATO Eimei

22　(2)　S50　2005/04/01

ISSN： 1341-7738
Morphological status of active mitochondria in vitrified bovine GV oocytes affects subsequent development.

Y Abe, N Aono, K Hara, H Matsumoto, H Sasada, E Sato

BIOLOGY OF REPRODUCTION　230-230　2005

ISSN： 0006-3363
Vitrification of germinal vesicle stage oocytes

Abe,Y, Aono,K, Hara,K, Matsumoto,H, Bakhtiyari,M, Sasada,H, Sato,E

Tohoku.J.Agri.Res.　55　(1)　45-50　2004

ISSN： 0040-8719
Nylon mesh is suitable for vitrification of large quantities of oocytes

Mstsumoto,H, Abe,Y, Hara,K, Sasada,H, Sato,E

REPROTECH　1　(3)　103-108　2004
Successful production of blastocysts following ultrarapid vitrification of germinal vesicle oocytes in mice by step-wise equilibrium.

N Aono, H Yoshida, Y Abe, K Hara, H Sasada, E Sato

FERTILITY AND STERILITY　80　S141-S142　2003/09

ISSN： 0015-0282

Show all ︎Show first 5

Research Projects 13

幹細胞に着目した高齢動物の精子産生能低下メカニズム

原健士朗, 鈴木伸之介

Offer Organization: 日本学術振興会

System: 科学研究費助成事業

Category: 基盤研究(B)

Institution: 東北大学

2025/04/01 - 2029/03/31
超寿命の家畜精子産生システム開発に資する精子幹細胞の性質解明

原健士朗

Offer Organization: 日本学術振興会

System: 科学研究費助成事業基盤研究(B)

Category: 基盤研究(B)

Institution: 東北大学

2021/04/01 - 2025/03/31

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2021年度は以下の３項目を実施し、精子幹細胞の遊走および細胞外基質分泌の制御機およびその種間共通性・相違性の検討を進めた。１）マウス未分化型精原細胞を対象として、既報のマイクロアレイもしくはRNAシークエンシングのデータセットの解析、細胞回収後のRT-PCRによる遺伝子発現解析、in situハイブリダイゼーションによるｍRNA局在解析、免疫組織化学によるタンパク質局在解析をそれぞれ実施し、未分化型精原細胞で発現する細胞外基質関連遺伝子・タンパク質の一部を明らかにした。本項目の一部は研究分担者とともに実施した。２）前項目で特定した遺伝子群のうち、その分子機能が精子幹細胞の移動において特に重要と予想された遺伝子１つについて細胞種特異的な遺伝子欠損マウスを作製し、2022年以降に同遺伝子の精子幹細胞における機能を検討するための技術基盤を確立した。なお、同遺伝子は全身性遺伝子欠損の場合に胎性致死となるため、Cre/LoxPシステムを応用した未分化型精原細胞特異的な遺伝子機能欠損マウスを作製した。３）前項目で着目した遺伝子・タンパク質について、精子幹細胞におけるその発現の種間差比較を行うため、成ウシ・ニワトリ精巣の採材・固定・包埋を行った。本項目の一部は研究分担者とともに実施した。以上、2021年の計画を予定通りに実施し、2022年以降の解析に必要な知見・技術が整った。本研究の推進により、実験・産業動物の精子幹細胞の特定・回収・培養に必要な幹細胞の細胞生物学的性質に関する知見が得られることが期待される。
精子の質的不均一性発生機構の解明

原健士朗, 盧尚建

Offer Organization: 日本学術振興会

System: 科学研究費助成事業

Category: 挑戦的研究(萌芽)

Institution: 東北大学

2021/07/09 - 2023/03/31

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2021年は、以下の解析を実施し、雄体内における精子の品質制御メカニズムの解析技術の開発を進めた。まず、精子の前駆細胞の膜上に時期特異的に蛍光標識を入れるためのCreERマウスの作製を進めた。既報のRNAシークエンス解析のデータセットから分化細胞で特異的に発現する遺伝子を１つ選択し、その発現を免疫組織化学染色で確認した後、同遺伝子のプロモータの下流にCreERを導入したベクターを作製後、培養下の胚性幹（ES）細胞内での相同組換えを行った。得られた組換えES細胞を正常型の胚盤胞にインジェクションし、現在までに毛への高い寄与が確認されたキメラを2系統作成し、F1を得るための交配試験を実施している。キメラマウスについて、生殖系列へも高い寄与が予想されるが、系統数が少なく、うまくF1が得られない場合に備え、追加のキメラ作製も検討中である。次に、Creによる組換えによって特異的に精子の前駆細胞を蛍光標識できるレポータマウスを検討した。具体的には、生殖細胞の細胞膜に蛍光タンパク質を取り込める既報のレポーターマウスを第一候補として導入した。上述のCreERマウスおよびレポーターマウスを掛け合わせることで、2022年に精子品質制御機構解析のモデル動物を開発する。なお、今後の解析に必要なイメージング技術の改良および内分泌影響の予備検討も実施した。以上の研究は、雄体内における精子の品質のばらつきの発生要因解析に資する技術基盤であり、この利活用を通じて野生動物や産業動物の雄繁殖能制御に資する成果が得られることが期待される。
精子形成を支える幹細胞のホメオスタシスと機能低下

吉田松生

System: AMED-CREST 全ライフコースを対象とした個体の機能低下機構の解明

2019/04 - 2023/03
精子産生における生殖細胞移動の役割

原健士朗

Offer Organization: JST

System: 創発的研究支援事業

2021/04 -
Elucidation of role of sperm stem cell motion

Hara Kenshiro

Offer Organization: Japan Society for the Promotion of Science

System: Grants-in-Aid for Scientific Research

Category: Grant-in-Aid for Challenging Research (Exploratory)

Institution: Tohoku University

2019/06 - 2021/03

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In this study, we aimed to understand the physiological roles of sperm stem cell motion on the basement membrane of mouse testis. Through the gene expression analyses, we revealed that sperm stem cells were expressed some extracellular matrix related genes which might contribute to maintain the homeostasis of basement membrane of seminiferous tubules. Further functional analysis will be needed to reveal the biological significance of these phenomenon.
精細管内フローと精子インテグリティ

原健士朗

Offer Organization: 日本学術振興会

System: 科学研究費助成事業新学術領域研究(研究領域提案型)

Category: 新学術領域研究(研究領域提案型)

Institution: 東北大学

2019/04 - 2021/03

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ウシで機能的精子を継続的に産生する体外培養系が構築できれば、種雄の個体飼育が不要になり、畜産業におけるウシ生産の高効率化、さらには動物資源生産の安定化が期待できる。本研究では、生体内の精子形成において重要と考えられる精細管内フローに着目して、ウシ精巣培養技術の改良を試みた。R3年度は、ウシ精細管を、前年度に作成したマイクロ流体デバイス内に静置し、内腔フローの再現を試みた。前年度に課題となっていた精細管の位置制御については微動インジェクターの使用により再現性良く制御することが可能となったが、流路内に圧を印加したところ、管内内腔の抵抗が予想よりも大きく、流れが管の外側に逃げ、内腔フローを再現することが出来なかった。この点については、その後も種々の検討を行ったものの、内腔フローを生じさせてその影響を検討するという当初の目的を達成することが出来なかった。一方、当初の計画になかった進捗としては、計画班が開発したPDMS製のカバーを用いて組織構造を薄い板状で培養することで、ウシ精巣片内の精細管の構造維持率および生殖細胞の生存性を向上させることができた。ウシ精巣培養におけるガスや栄養環境の均一化の重要性を示すものであったが、減数分裂以降の分化の再現は依然として課題として残った。今後、ウシ精巣培養下で生殖細胞の分化を阻害する要因を探索し、これを取り除くことで、ウシ体外精子形成技術の改良を進める予定である。
雄体内における精子移動に着目した精子の質的ヘテロジェネイティ生成機構

原健士朗

Offer Organization: 住友財団

System: 基礎科学研究助成

2019/11 - 2020/10
Investigating the mechanisms for GSC regulation in mouse gametogenesis

YOSHIDA Shosei

Offer Organization: Japan Society for the Promotion of Science

System: Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)

Category: Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)

Institution: National Institute for Basic Biology

2015/06 - 2018/03

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Continual production of numerous sperm in the testes ensured the continuity of life. Here, spermatogenic stem cells play essential roles through giving rise to both the sperm-producing differentiating cells and the self-renewing stem cells, in a well balance manner. This study has contributed toward the understanding of the nature of spermatogenic stem cells by proposing a novel model for the stem cell dynamics, discovering a crucial change in the status of genome DNA, discovery of novel molecular mechanisms making the selfrenewal and differentiation compatible in uniform tissue environment, as well as investigating the mechanisms underlying the stem cell number regulation and post-transplantation repopulation.
幹細胞の柔軟性を生かした抗老化システムの解明

原健士朗

Offer Organization: 日本学術振興会

System: 科学研究費助成事業新学術領域研究(研究領域提案型)

Category: 新学術領域研究(研究領域提案型)

Institution: 東北大学

2015/04 - 2017/03

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本研究は、組織幹細胞の中でも特に老化しにくいとされる精子幹細胞をモデルとして、生理的な加齢条件下での、マウス精子幹細胞の挙動の加齢変化を明らかにすることを目的とした。具体的には、①幹細胞の不完全分裂（不完全な細胞質分裂によって細胞分裂後も各細胞が連結したままとなる分裂様式）と合胞体（不完全分裂の結果生じる複数の細胞が連結した構造体）の断片化（細胞間架橋が千切れて合胞体が短くなる現象）の頻度、および②幹細胞から分化した細胞の脱分化の頻度、の２項目の加齢変化を検証する事を計画した。昨年度（H27）は、①を検討し、マウス精子幹細胞の合胞体の断片化頻度が加齢に伴って上昇することを示唆する結果を得た。この加齢変化は、幹細胞として機能する合胞体の数を増やす役割を果たすため、加齢に伴う精子数減少を食い止める新現の抗老化現象となる可能性が考えられた。一方、本年度(H28)は、②を検証するため、分化細胞の脱分化による幹細胞の補充が加齢に伴って増加し、精子形成維持に寄与しているという仮説をたて、解析を進めた。若齢（3か月齢）および加齢（14～19か月）のNgn3-CreER;CAG-CAT-GFPマウスを用いて、Ngn3陽性の分化細胞の運命を数日～数十日間追跡し、GFRα1陽性に逆戻りする頻度を解析した結果、加齢に伴った逆戻り頻度の変化は認められず、仮説は棄却された。以上、H27,H28の研究から、精子幹細胞の合胞体断片化頻度の加齢変化が明らかになった。今後、加齢変化を生み出す分子メカニズムの解明と同機構破綻の影響の解明が期待される。
Elucidation of patterns and regulatory mechanism of undifferentiated spermatogonial motion in mouse testis

Hara Kenshiro

Offer Organization: Japan Society for the Promotion of Science

System: Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

Category: Grant-in-Aid for Scientific Research (C)

2014/04 - 2017/03

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The purpose of this study is to understand motion patterns and regulatory mechanisms of stem and differentiating cells in mouse seprmatogenesis. In this study, we found that stem cells move actively and randomly, while differentiating cells are less motile. We also found that heterogeneous motion is likely to be generated by two mechanisms: syncytial formation (unique biological event in spermatogenesis) and cell state-dependent motion property. Future analysis for molecular mechanisms of stem/differentiating cell motion and biological significance of stem cell motion will be needed. During this support, we published a paper in Cell Stem Cell (Hara etal., 2014).
Identification of stem cells in mouse spermatogenesis

HARA Kenshiro

Offer Organization: Japan Society for the Promotion of Science

System: Grants-in-Aid for Scientific Research Grant-in-Aid for Young Scientists (B)

Category: Grant-in-Aid for Young Scientists (B)

Institution: National Institute for Basic Biology

2010 - 2012

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To identify stem cells in mouse spermatogenesis, I analysed the fate behaviour of GFRα1-expressing spermatogonia and their progeny at single-cell resolution by combining in vivo live-imaging with pulse-labeling studies. Data suggest that the morphologically heterogeneous population of GFRα1-expressing spermatogonia together comprise a stem cell pool
腸管形成不全モデル動物を利用した始原生殖細胞の移動・分化メカニズムの解析

原健士朗

Offer Organization: 日本学術振興会

System: 科学研究費助成事業特別研究員奨励費

Category: 特別研究員奨励費

Institution: 東京大学

2006 - 2008

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本研究では、PGCsと胚性内胚葉の相互作用を明らかにするため、Geneticalな体細胞組織の移動阻害実験を計画した。Sox(S__-ry-related HMG b<ox>___-)サブグループFに属するSox17遺伝子が胚性内胚葉に発現しており、その欠損マウス胚では胚性内胚葉の細胞数が減少することが報告されている。つまり、同マウス胚はPGCsの移動の足場である胚性内胚葉の形成不全モデルマウス胚といえる。そこで、胚性内胚葉形成不全がPGCsの移動・分化に及ぼす影響を明らかにすることを目的とし、Sox17欠損マウス胚におけるPGCsの移動・分化を解析した。 Sox17欠損マウスの解析結果から、哺乳類の生殖細胞は形成直後に胚性内胚葉組織に移入すると、胚性内胚葉によって自律的な運動能を抑制され、胚性内胚葉の分化とそれに伴う形態形成の力を利用して後腸まで移動することが明らかとなった。同研究成果は、連綿と続く哺乳類の生命の連続性を保証するメカニズムとして重要なものであるばかりでなく、しばしばその性質から"脳"に例えられる腸管の発生過程におけるもっとも初期の役割を明確に示したという点においても極めて興味深いものである。

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