The Experts below are selected from a list of 11991 Experts worldwide ranked by ideXlab platform
Ralph L. Brinster - One of the best experts on this subject based on the ideXlab platform.
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Transplantation of testis germinal cells into mouse Seminiferous Tubules.
The International Journal of Developmental Biology, 2003Co-Authors: Takehiko Ogawa, Juan Aréchaga, Mary R. Avarbock, Ralph L. BrinsterAbstract:In the adult male, germ cell differentiation takes place in the Seminiferous Tubules of the testis by a complex, highly organized and very efficient process. A population of diploid stem-cell spermatogonia that lie on the basement membrane of the tubule continuously undergoes self-renewal and produces progeny cells, which initiate the process of cellular differentiation to generate mature spermatozoa. Each testis contains many Seminiferous Tubules, which are connected at both ends to a collecting system called the rete testis. The mature spermatozoa pass from the Tubules into the rete and are then carried through efferent ducts to the epididymis for final maturation before they are ready to fertilize an egg. In previous studies, we have demonstrated that donor testis cells collected from a fertile mouse are able to generate spermatogenesis when transplanted to the Seminiferous Tubules of an infertile male. The spermatozoa produced by the recipient from the donor-derived spermatogonial stem cells are able to fertilize eggs and produce progeny carrying the donor male haplotype. Furthermore, donor testis stem cells from a rat will generate normal rat spermatozoa following transplantation to a mouse testis. The spermatogonial transplantation technique is clearly valuable and applicable to many species, but it is difficult. Therefore, several procedures to introduce donor cells into the Seminiferous Tubules of a recipient have been developed using the mouse as a model, and they are described here in detail. The results indicate that microinjection of cell suspensions into the Seminiferous Tubules, efferent ducts or rete testis are equally effective in generating donor cell-derived spermatogenesis in recipients. Each approach is likely to be useful for different experimental purposes in a variety of species.
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computer assisted image analysis to assess colonization of recipient Seminiferous Tubules by spermatogonial stem cells from transgenic donor mice
Molecular Reproduction and Development, 1999Co-Authors: Ina Dobrinski, Takehiko Ogawa, Mary R. Avarbock, Ralph L. BrinsterAbstract:Transplantation of spermatogonial stem cells from fertile, transgenic donor mice to the testes of infertile recipients provides a unique system to study the biology of spermatogonial stem cells. To facilitate the investigation of treatment effects on colonization efficiency an analysis system was needed to quantify colonization of recipient mouse Seminiferous Tubules by donor stem cell-derived spermatogenesis. In this study, a computer-assisted morphometry system was developed and validated to analyze large numbers of samples. Donor spermatogenesis in recipient testes is identified by blue staining of donor-derived spermatogenic cells expressing the E. coli lacZ structural gene. Images of Seminiferous Tubules from recipient testes collected three months after spermatogonial transplantation are captured, and stained Seminiferous Tubules containing donor-derived spermatogenesis are selected for measurement based on their color by color thresholding. Colonization is measured as number, area, and length of stained Tubules. Interactive, operator-controlled color selection and sample preparation accounted for less than 10% variability for all collected parameters. Using this system, the relationship between number of transplanted cells and colonization efficiency was investigated. Transplantation of 10(4) cells per testis only rarely resulted in colonization, whereas after transplantation of 10(5) and 10(6) cells per testis the extent of donor-derived spermatogenesis was directly related to the number of transplanted donor cells. It appears that about 10% of transplanted spermatogonial stem cells result in colony formation in the recipient testis. The present study establishes a rapid, repeatable, semi-interactive morphometry system to investigate treatment effects on colonization efficiency after spermatogonial transplantation in the mouse.
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Ultrastructural Observations of Spermatogenesis Following Transplantation of Rat Testis Cells Into Mouse Seminiferous Tubules
Journal of andrology, 1996Co-Authors: Lonnie D. Russell, Ralph L. BrinsterAbstract:The testes of busulfan-treated immunodeficient mice receiving Seminiferous tubule injections of testis cells from rats were examined by light and electron microscopy. The presence of active rat spermatogenesis was verified by criteria that are known to characterize spermatogenic cells of this species. In addition, spermatogenesis from the mouse was identified as taking place in some Seminiferous Tubules as the result of reinitiation of spermatogenesis after busulfan treatment. Rat spermatogenesis in mouse Seminiferous Tubules showed the generally recognized associations of cells known to characterize stages of spermatogenesis of the rat. The Sertoli cells associated with rat spermatogenesis were identified ultrastructurally as being of mouse origin. Thus, rat spermatogenesis, which has a cycle length that is 50% longer than mouse spermatogenesis, can proceed among mouse Sertoli cells, which supposedly exert much shorter cyclic influences in concert with mouse germ cell development. Studies are needed to determine if the timing of rat spermatogenesis is controlled by the germ cells or the Sertoli cells. These observations are considered preliminary since a thorough study of somatic-germ cell relationships was not undertaken. It is concluded that a mouse Sertoli cell in the environment provided by the mouse testis can produce both mouse and rat gametes.
Renee Reijo A Pera - One of the best experts on this subject based on the ideXlab platform.
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fate of induced pluripotent stem cells following transplantation to murine Seminiferous Tubules
Human Molecular Genetics, 2014Co-Authors: Jens Durruthy Durruthy, Cyril Ramathal, Meena Sukhwani, Kyle E Orwig, Fang Fang, Jun Cui, Renee Reijo A PeraAbstract:Studies of human germ cell development are limited in large part by inaccessibility of germ cells during development. Moreover, although several studies have reported differentiation of mouse and human germ cells from pluripotent stem cells (PSCs) in vitro, differentiation of human germ cells from PSCs in vivo has not been reported. Here, we tested whether mRNA reprogramming in combination with xeno-transplantation may provide a viable system to probe the genetics of human germ cell development via use of induced pluripotent stem cells (iPSCs). For this purpose, we derived integration-free iPSCs via mRNA-based reprogramming with OCT3/4, SOX2, KLF4 and cMYC alone (OSKM) or in combination with the germ cell-specific mRNA, VASA (OSKMV). All iPSC lines met classic criteria of pluripotency. Moreover, global gene expression profiling did not distinguish large differences between undifferentiated OSKM and OSKMV iPSCs; however, some differences were observed in expression of pluripotency factors and germ cell-specific genes, and in epigenetic profiles and in vitro differentiation studies. In contrast, transplantation of undifferentiated iPSCs directly into the Seminiferous Tubules of germ cell-depleted immunodeficient mice revealed divergent fates of iPSCs produced with different factors. Transplantation resulted in morphologically and immunohistochemically recognizable germ cells in vivo, particularly in the case of OSKMV cells. Significantly, OSKMV cells also did not form tumors while OSKM cells that remained outside the Seminiferous tubule proliferated extensively and formed tumors. Results indicate that mRNA reprogramming in combination with transplantation may contribute to tools for genetic analysis of human germ cell development.
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fate of ipscs derived from azoospermic and fertile men following xenotransplantation to murine Seminiferous Tubules
Cell Reports, 2014Co-Authors: Cyril Ramathal, Jens Durruthydurruthy, Meena Sukhwani, Joy E Arakaki, Paul J Turek, Kyle E Orwig, Renee Reijo A PeraAbstract:Historically, spontaneous deletions and insertions have provided means to probe germline developmental genetics in Drosophila, mouse and other species. Here, induced pluripotent stem cell (iPSC) lines were derived from infertile men with deletions that encompass three Y chromosome azoospermia factor (AZF) regions and are associated with production of few or no sperm but normal somatic development. AZF-deleted iPSC lines were compromised in germ cell development in vitro. Undifferentiated iPSCs transplanted directly into murine Seminiferous Tubules differentiated extensively to germ-cell-like cells (GCLCs) that localized near the basement membrane, demonstrated morphology indistinguishable from fetal germ cells, and expressed germ-cell-specific proteins diagnostic of primordial germ cells. Alternatively, all iPSCs that exited Tubules formed primitive tumors. iPSCs with AZF deletions produced significantly fewer GCLCs in vivo with distinct defects in gene expression. Findings indicate that xenotransplantation of human iPSCs directs germ cell differentiation in a manner dependent on donor genetic status.
Takehiko Ogawa - One of the best experts on this subject based on the ideXlab platform.
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in vitro reconstruction of mouse Seminiferous Tubules supporting germ cell differentiation
Biology of Reproduction, 2013Co-Authors: Tetsuhiro Yokonishi, Yoshinobu Kubota, Takuya Sato, Kumiko Katagiri, Mitsuru Komeya, Takehiko OgawaAbstract:It is known that cells of testis tissues in fetal or neonatal periods have the ability to reconstruct the testicular architecture even after dissociation into single cells. This ability, however, has not been demonstrated effectively in vitro. In our present study, we succeeded in reconstructing Seminiferous Tubules in vitro which supported spermatogenesis to meiotic phase. Testis cells of neonatal mice were dissociated enzymatically into single cells. The cells formed aggregates in suspension culture and were transferred to the surface of agarose gel to continue the culture with a gas-liquid interphase method, where a tubular architecture gradually developed during the following 2 weeks. Immunohistological examination confirmed Sertoli cells forming Tubules and germ cells inside. With testis tissues of Acr-GFP transgenic mice, whose germ cells express GFP during meiosis, cell aggregates formed a tubular structure and showed GFP expressions in their reconstructed tissues. Meiotic figures were also confirmed by regular histology and immunohistochemistry. In addition, we mixed cell lines of spermagonial stem cells (GS cells) into the testis cell suspension, and found the incorporation of GS cells in the Tubules in reconstructed tissues. When GS cells derived from Acr-GFP transgenic mice were used, GFP expression was observed, indicating that the spermatogenesis of GS cells was proceeding up to the meiotic phase. This in vitro reconstruction technique will be a useful method for the study of testis organogenesis and spermatogenesis.
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Transplantation of testis germinal cells into mouse Seminiferous Tubules.
The International Journal of Developmental Biology, 2003Co-Authors: Takehiko Ogawa, Juan Aréchaga, Mary R. Avarbock, Ralph L. BrinsterAbstract:In the adult male, germ cell differentiation takes place in the Seminiferous Tubules of the testis by a complex, highly organized and very efficient process. A population of diploid stem-cell spermatogonia that lie on the basement membrane of the tubule continuously undergoes self-renewal and produces progeny cells, which initiate the process of cellular differentiation to generate mature spermatozoa. Each testis contains many Seminiferous Tubules, which are connected at both ends to a collecting system called the rete testis. The mature spermatozoa pass from the Tubules into the rete and are then carried through efferent ducts to the epididymis for final maturation before they are ready to fertilize an egg. In previous studies, we have demonstrated that donor testis cells collected from a fertile mouse are able to generate spermatogenesis when transplanted to the Seminiferous Tubules of an infertile male. The spermatozoa produced by the recipient from the donor-derived spermatogonial stem cells are able to fertilize eggs and produce progeny carrying the donor male haplotype. Furthermore, donor testis stem cells from a rat will generate normal rat spermatozoa following transplantation to a mouse testis. The spermatogonial transplantation technique is clearly valuable and applicable to many species, but it is difficult. Therefore, several procedures to introduce donor cells into the Seminiferous Tubules of a recipient have been developed using the mouse as a model, and they are described here in detail. The results indicate that microinjection of cell suspensions into the Seminiferous Tubules, efferent ducts or rete testis are equally effective in generating donor cell-derived spermatogenesis in recipients. Each approach is likely to be useful for different experimental purposes in a variety of species.
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computer assisted image analysis to assess colonization of recipient Seminiferous Tubules by spermatogonial stem cells from transgenic donor mice
Molecular Reproduction and Development, 1999Co-Authors: Ina Dobrinski, Takehiko Ogawa, Mary R. Avarbock, Ralph L. BrinsterAbstract:Transplantation of spermatogonial stem cells from fertile, transgenic donor mice to the testes of infertile recipients provides a unique system to study the biology of spermatogonial stem cells. To facilitate the investigation of treatment effects on colonization efficiency an analysis system was needed to quantify colonization of recipient mouse Seminiferous Tubules by donor stem cell-derived spermatogenesis. In this study, a computer-assisted morphometry system was developed and validated to analyze large numbers of samples. Donor spermatogenesis in recipient testes is identified by blue staining of donor-derived spermatogenic cells expressing the E. coli lacZ structural gene. Images of Seminiferous Tubules from recipient testes collected three months after spermatogonial transplantation are captured, and stained Seminiferous Tubules containing donor-derived spermatogenesis are selected for measurement based on their color by color thresholding. Colonization is measured as number, area, and length of stained Tubules. Interactive, operator-controlled color selection and sample preparation accounted for less than 10% variability for all collected parameters. Using this system, the relationship between number of transplanted cells and colonization efficiency was investigated. Transplantation of 10(4) cells per testis only rarely resulted in colonization, whereas after transplantation of 10(5) and 10(6) cells per testis the extent of donor-derived spermatogenesis was directly related to the number of transplanted donor cells. It appears that about 10% of transplanted spermatogonial stem cells result in colony formation in the recipient testis. The present study establishes a rapid, repeatable, semi-interactive morphometry system to investigate treatment effects on colonization efficiency after spermatogonial transplantation in the mouse.
Lonnie D. Russell - One of the best experts on this subject based on the ideXlab platform.
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non random distribution of spermatogonia in rats evidence of niches in the Seminiferous Tubules
Reproduction, 2003Co-Authors: Helio Chiarinigarcia, A M Raymer, Lonnie D. RussellAbstract:The relationships and distribution of spermatogonia were studied as a function of the stage of the Seminiferous epithelium cycle in rats. Primitive spermatogonia in the mouse are located along regions of the basal lamina that face the interstitium. Before studying the distribution of spermatogonia in rats, it was necessary to characterize the various types of spermatogonia, as recently performed for mice. The Strauss’ linear index (Li) selectivity method was then used and spermatogonia of the Asingle (As )t o Aaligned (Aal) lineage were preferentially found to be located in regions opposing the interstitium at stages V, VII and IX of the spermatogenic cycle. Because relatively little tubuleto-tubule contact occurs in rats, the aim of this study was to determine whether tubule-to-tubule contact or tubule proximity (or alternatively, the amount of interstitium) was an important factor in spermatogonial position. In this regard, another method (tubule proximity) was devised to determine spermatogonial position that accounted for the presence of adjacent Tubules. This method showed that the position of Tubules, rather than tubule contact, was more accurate than the Li method in determining the location of spermatogonia in the rat. The results also showed a non-random distribution of spermatogonia resembling that of the mouse, and that tubule-to-tubule contact is not essential for the positioning of spermatogonia. In conclusion, the results of this study strongly indicate that the most primitive type A spermatogonia (As ,A paired and Aal )i n rats are present in niches located in those areas of the Seminiferous Tubules that border the interstitial tissue.
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juvenile spermatogonial depletion jsd mutant Seminiferous Tubules are capable of supporting transplanted spermatogenesis
Biology of Reproduction, 2000Co-Authors: Holly L Boettgertong, Michael D. Griswold, Lonnie D. Russell, Daniel S Johnston, Colin E BishopAbstract:In mice, the juvenile spermatogonial depletion (jsd) mutation results in a single wave of spermatogenesis followed by failure of type A spermatogonial stem cells to repopulate the testis, rendering male animals sterile. It is not clear whether the defect in jsd resides in a failure of the somatic component to support spermatogenesis or in a failure that is intrinsic to the mutant’s germ cells. To determine if the jsd intratesticular environment is capable of supporting spermatogenesis, germ cell transplantation experiments were performed in which C57BL/6 ROSA germ cells were transplanted into jsd recipients. To determine if jsd spermatogonia are able to develop in a permissive Seminiferous environment, jsd germ cells were transplanted into W/W v and busulfan-treated C57BL/6 animals. The data demonstrate that up to 7 mo after transplantation of normal germ cells, jsd Seminiferous Tubules are capable of supporting spermatogenesis. In contrast, when jsd germ cells were transplanted into busulfantreated C57BL/6 testis, or into testis of W/W v mice, no jsd-derived spermatogenesis was observed. The data support the hypothesis that the jsd phenotype is due to a defect in the germ cells themselves, and not in the intratubular environment. gametogenesis, processes, spermatogenesis, testes
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Ultrastructural Observations of Spermatogenesis Following Transplantation of Rat Testis Cells Into Mouse Seminiferous Tubules
Journal of andrology, 1996Co-Authors: Lonnie D. Russell, Ralph L. BrinsterAbstract:The testes of busulfan-treated immunodeficient mice receiving Seminiferous tubule injections of testis cells from rats were examined by light and electron microscopy. The presence of active rat spermatogenesis was verified by criteria that are known to characterize spermatogenic cells of this species. In addition, spermatogenesis from the mouse was identified as taking place in some Seminiferous Tubules as the result of reinitiation of spermatogenesis after busulfan treatment. Rat spermatogenesis in mouse Seminiferous Tubules showed the generally recognized associations of cells known to characterize stages of spermatogenesis of the rat. The Sertoli cells associated with rat spermatogenesis were identified ultrastructurally as being of mouse origin. Thus, rat spermatogenesis, which has a cycle length that is 50% longer than mouse spermatogenesis, can proceed among mouse Sertoli cells, which supposedly exert much shorter cyclic influences in concert with mouse germ cell development. Studies are needed to determine if the timing of rat spermatogenesis is controlled by the germ cells or the Sertoli cells. These observations are considered preliminary since a thorough study of somatic-germ cell relationships was not undertaken. It is concluded that a mouse Sertoli cell in the environment provided by the mouse testis can produce both mouse and rat gametes.
Ping Liu - One of the best experts on this subject based on the ideXlab platform.
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up regulation of heme oxygenase 1 expression modulates reactive oxygen species level during the cryopreservation of human Seminiferous Tubules
Fertility and Sterility, 2014Co-Authors: Wenhao Tang, Ping Liu, Jie Yan, Tianren Wang, Xi Xia, Xinjie Zhuang, Kai HongAbstract:Objective To study the effect of freezing techniques and to optimize a method for trace amounts of testicular spermatozoa from biopsed Seminiferous Tubules. The level of reactive oxygen species (ROS) and the gene expression of heme oxygenase-1 was evaluated. Design Prospective experimental study. Setting University-based laboratory. Patient(s) Eighteen adults with male fator infertility underwent testicular biopsy surgery. Intervention(s) Seminiferous tubular fragments from each man were evenly allocated to three groups: fresh control, slow freezing, and vitrifiaction groups. The morphology and ROS levels before and after freezing were evaluated for Seminiferous tubular fragments. The expression of heme oxygenase-1 (HO-1) at both the transcriptional and protein levels was determined. Main Outcome Measure(s) The morphology was analyzed by light microscopy. The ROS levels were measured with ELISA. The proliferation and differentiation were evaluated by immunohistochemistry, and the expression of HO-1 was evaluated using a real-time polymerase chain reaction (PCR) and Western blotting. Result(s) Decreased ROS levels and increased HO-1 expression at the transcriptional and protein levels were observed after thawing the human Seminiferous Tubules. The ROS level was negatively correlated with HO-1 expression. Slow freezing was more effective than vitrification in terms of HO-1 up-regulation and ROS alteration. Conclusion(s) Based on our study, the slow freezing technique was more effective compared with the vitrification method.
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derivation of male germ cells from induced pluripotent stem cells in vitro and in reconstituted Seminiferous Tubules
Cell Proliferation, 2012Co-Authors: Shiping Yang, Xiangjiang Guo, Ruhui Tian, Can Sun, Yinjie Zhu, Ping Liu, Shasha Zou, Yuhua HuangAbstract:Objectives Previous studies have demonstrated that mouse- and human-induced pluripotent stem (iPS) cells can differentiate into primordial germ cells in vitro. However, up to now it is not known whether iPS cells would be able to differentiate into male germ cells in vivo. The aim of this study was to explore differentiation potential of iPS cells to male germ cells in vitro and in vivo. Materials and methods In this study, approaches using in vitro retinoic acid induction and in vivo ectopic transplantation were combined to induce iPS cells to become male germ cells. Results RT-PCR showed that expression of pre-meiotic and meiotic germ cell-specific genes was enhanced in iPS cell-derived embryoid bodies (EBs) compared to mRNA transcripts of iPS cells. Immunofluorescence analysis revealed that iPS cell-derived EBs positively expressed germ-cell markers VASA, c-Kit and SCP3. Furthermore, iPS cell-derived cells dissociated from EBs were injected with testicular cells into dorsal skin of mice. Histological examination showed that iPS cell-derived cells could reconstitute Seminiferous Tubules, and meanwhile, iPS cell-derived germ cells could settle at basement membranes of reconstituted Tubules. Conclusion Our results suggest that iPS cells are able to differentiate into male germ cells in vitro and that reconstituted Seminiferous Tubules may provide a functional niche for exogenous iPS cell-derived male germ cells. Derivation of male germ cells from iPS cells has potential application for treating male infertility and provides an ideal platform for elucidating molecular mechanisms of male germ-cell development.
