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ABSTRACT: Tiger puffer Takifugu rubripes is one of the most valuable fish species in Japan; however, there has not been much progress in their selective breeding until recently despite their potential in aquaculture. Their long generation time and the large body size of their broodstock make breeding difficult. Recently, we made a surrogate broodstock, which produced gametes of different species in salmonids. Therefore, by using closely related recipients, which have small body sizes and short generation times, it is possible to accelerate breeding of the tiger puffer. Thus, we considered the grass puffer Takifugu niphobles, which has a short generation time and a small maturation size, as a potential recipient for gamete production of the tiger puffer. Furthermore, if sterile triploid individuals are used as recipients, the resulting surrogate broodstock would produce only donor-derived gametes. Therefore, we examined conditions for inducing triploidy by suppressing meiosis II to retain the second polar body in grass puffer. We found that cold shock treatment, which is 5°C for 30 min starting from 5 min after fertilization, is optimal to obtain high triploidization and hatching rates. Although the resulting triploid grass puffers produced small amounts of gametes in both sexes, the offspring derived from the gametes could not live for over 3 days. Furthermore, we found that triploid grass puffer showed normal plasma sex steroid levels compared with diploids. These are important characteristics of triploid grass puffer as surrogate recipients used for germ cell transplantation.
Marine Biotechnology 07/2012; · 3.43 Impact Factor
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ABSTRACT: In spermatogonial transplantation using Pacific bluefin tuna Thunnus orientalis as a donor, enrichment of spermatogonia (SG) is expected to facilitate high colonization efficiency. Although it is desirable to establish a bluefin tuna SG enrichment procedure using cell-surface markers, a germ cell-specific cell-surface marker has not been identified to date. We previously found that Ly75 is a mitotic germ cell-specific cell-surface marker in rainbow trout, and that its amino-acid sequences are highly conserved in various teleosts. Thus, the ly75 gene is an excellent candidate cell-surface marker of SG in bluefin tuna. In this study, the bluefin tuna ly75 homolog was cloned and characterized for further use as a germ cell-specific cell-surface marker. In adult tissues, high levels of ly75 transcripts were detected in the liver, pyloric caeca, and testis. In situ hybridization analyses showed that ly75 mRNA was predominantly localized in type-A spermatogonia (A-SG), including single A-SG that contain transplantable germ cells. In contrast, ly75 mRNA was not detected in spermatocytes, spermatids, or gonadal somatic cells in testis. The expression profiles of Ly75 protein were similar to those of the mRNA. Therefore, Ly75 is appropriate for use as a cell-surface marker of SG in bluefin tuna.
Fisheries Science 05/2012; 78(4):791-800. · 0.94 Impact Factor
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ABSTRACT: In spermatogonial transplantation using Pacific bluefin tuna Thunnus orientalis as a donor, enrichment of spermatogonia (SG) is expected to facilitate high coloni-zation efficiency. Although it is desirable to establish a bluefin tuna SG enrichment procedure using cell-surface markers, a germ cell-specific cell-surface marker has not been identified to date. We previously found that Ly75 is a mitotic germ cell-specific cell-surface marker in rainbow trout, and that its amino-acid sequences are highly con-served in various teleosts. Thus, the ly75 gene is an excellent candidate cell-surface marker of SG in bluefin tuna. In this study, the bluefin tuna ly75 homolog was cloned and characterized for further use as a germ cell-specific cell-surface marker. In adult tissues, high levels of ly75 transcripts were detected in the liver, pyloric caeca, and testis. In situ hybridization analyses showed that ly75 mRNA was predominantly localized in type-A spermato-gonia (A-SG), including single A-SG that contain trans-plantable germ cells. In contrast, ly75 mRNA was not detected in spermatocytes, spermatids, or gonadal somatic cells in testis. The expression profiles of Ly75 protein were similar to those of the mRNA. Therefore, Ly75 is appro-priate for use as a cell-surface marker of SG in bluefin tuna.
Fisheries Science 05/2012; 78(4):791-800. · 0.94 Impact Factor
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ABSTRACT: Recently, we developed an intraspecies spermatogonial transplantation technique in a pelagic egg spawning marine teleost,
nibe croaker Nibea mitsukurii. Nibe croaker is an ideal candidate recipient for spermatogonial transplantation since it has a short generation time and
small body size. In the present study, yellowtail Seriola quinqueradiata spermatogonia were transplanted into nibe croaker larvae, and the behavior of transplanted spermatogonia in recipient gonads
was observed. Three weeks post-transplantation, yellowtail spermatogonia were incorporated into the gonads of 72 out of 88
recipients. An antiproliferating cell nuclear antigen was detected in incorporated yellowtail spermatogonia, suggesting that
the xenogenic germ cells were proliferating in recipient gonads. Yellowtail vasa-positive spermatogonia survived for 11months after transplantation in the gonads of recipient fish. Thus, we showed that
the microenvironment in nibe croaker gonads can support the colonization, proliferation, and survival of germ cells derived
from a different taxonomic family.
KeywordsXenogenic transplantation–Spermatogonia–Marine teleost–
Vasa
Fisheries Science 04/2012; 77(1):69-77. · 0.94 Impact Factor
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ABSTRACT: We identified fatty acid desaturase (fads)-like and elongase (elovl)-like genes from nibe croaker to better understand the molecular basis of n-3 highly unsaturated fatty acid metabolism in
marine fish. Phylogenetic analysis revealed that the fads-like and elovl-like genes were classified into the fads6 and elovl5 groups, respectively. We investigated the effects of various levels of docosahexaenoic acid (DHA)-enriched live feed, Artemia nauplii, on larval growth, survival, and fads-like and elovl-like gene expression. After a 15-day feed trial, total length, body weight, and survival were all significantly improved
by the supplementation of Artemia with DHA. This result indicates that nibe croaker cannot endogenously produce enough DHA. Furthermore, the fads-like gene transcripts in larvae fed on oleic acid-enriched Artemia were significantly higher than those in larvae on 100% DHA-enriched Artemia. In contrast, no significant differences were observed in the transcript levels of the elovl-like gene. These data indicate that the fads6-like gene was controlled by negative feedback from the quantity of DHA stored in the larval body. These results have implications
for the functionality of the fads-like gene in nibe croaker.
Keywords
Artemia
-Docosahexaenoic acid-n-3 Highly unsaturated fatty acid-Fatty acid desaturase-Fatty acid elongase-
Nibea mitsukurii
Fisheries Science 04/2012; 76(3):463-472. · 0.94 Impact Factor
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Tetsuro Morita,
Naoki Kumakura,
Kagayaki Morishima,
Toru Mitsuboshi,
Masashi Ishida,
Takashi Hara,
Satomi Kudo,
Misako Miwa,
Shoko Ihara,
Kentaro Higuchi, Yutaka Takeuchi,
Goro Yoshizaki
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ABSTRACT: Although the yellowtail (Seriola quinqueradiata) is the fish most commonly farmed in Japan, breeding of this species has not yet started. This is primarily due to the lack of sufficiently sophisticated methods for manipulating gametogenesis, which makes it difficult to collect gametes from specific dams and sires. If it were possible to produce large numbers of surrogate fish by transplanting germ cells isolated from donor individuals harboring desirable genetic traits, then the probability of acquiring gametes carrying the donor-derived haplotype would increase, and breeding programs involving this species might increase as a result. As a first step, we established a method for the allogeneic transplantation of yellowtail spermatogonia and the production of donor-derived offspring. Donor cells were collected from immature (10-month-old) yellowtail males with testes containing abundant type A spermatogonia, labeled with PKH26 fluorescent dye, and transferred into the peritoneal cavities of 8-day-old larvae. Fluorescence observation at 28 days post-transplantation revealed that PKH26-labeled cells were incorporated into recipients' gonads. To assess whether donor-derived spermatogonia could differentiate into functional gametes in the allogeneic recipient gonads, gametes collected from nine male and four female adult recipients were fertilized with wild-type eggs and milt. Analysis of microsatellite DNA markers confirmed that some of the first filial (F(1)) offspring were derived from donor fish, with the average contribution of donor-derived F(1) offspring being 66% and the maximum reaching 99%. These findings confirmed that our method was effective for transplanting yellowtail spermatogonia into allogeneic larvae to produce donor-derived offspring.
Biology of Reproduction 03/2012; 86(6):176. · 4.01 Impact Factor
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ABSTRACT: The transplantation of germ cells is a powerful tool both for studying their development and for reproductive biotechnology. An intraperitoneal germ cell transplantation system was recently developed for use in several teleost species. Donor germ cells transplanted into the peritoneal cavity of hatchlings migrated toward and were incorporated into the recipient's genital ridges, where they underwent gametogenesis. Among male germ cells, only type A spermatogonia were capable of colonizing the recipient gonads, unlike those at more advanced stages. The enrichment of type A spermatogonia is therefore important to achieve efficient donor-cell incorporation and subsequent donor-derived gametogenesis. Here we established a simple and rapid system of isolation and enrichment for fish type A spermatogonia, using flow cytometry. Type A spermatogonia were found to have distinctive forward and side light scatter properties compared to that with other types of testicular cell. Based on these characteristics, we were able to isolate and enrich type A spermatogonia by using flow cytometry. After intraperitoneal transplantation, the enriched type A spermatogonia could be successfully incorporated into the recipient genital ridges. This flow cytometry approach using forward and side light scatter was also found to be applicable to other salmonid and sciaenid species, suggesting that it could be a powerful tool for isolating and enriching transplantable type A spermatogonia in a wide range of teleosts. We expect this method to contribute significantly to germ cell biology and biotechnology.
Biology of Reproduction 01/2012; 86(4):107. · 4.01 Impact Factor
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ABSTRACT: Recent progress in genome-based breeding has created various fish strains carrying desirable genetic traits; however, methods for the long-term preservation of their genetic resources have not yet been developed, mainly due to the lack of cryopreservation techniques for fish eggs and embryos. Recently, we established an alternative cryopreservation technique for fish spermatogonia using a slow-freezing method. Furthermore, we developed a transplantation system to produce functional eggs and sperm derived from spermatogonia. Spermatogonia isolated from the testes of vasa-green fluorescent protein (Gfp) transgenic rainbow trout (Oncorhynchus mykiss) were transplanted into the peritoneal cavity of triploid masu salmon (Oncorhynchus masou) hatchlings of both genders. The transplanted trout spermatogonia migrated towards the gonadal anlagen of the recipient salmon, into which they were subsequently incorporated. We confirmed that the donor-derived spermatogonia resumed gametogenesis, and produced sperm and eggs in male and female recipient salmon, respectively. Fertilization of the resultant eggs and sperm produced only rainbow trout in the first filial (F₁) generation, suggesting that the sterile triploid recipient salmon produced functional eggs and sperm derived from the trout donors. A combination of spermatogonial transplantation and cryopreservation could be a powerful tool for preserving valuable fish strains with desirable genetic traits and endangered species.
Comparative Biochemistry and Physiology Part D Genomics and Proteomics 03/2011; 6(1):55-61. · 1.72 Impact Factor
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ABSTRACT: In mammals, several cell surface molecular markers have been characterized in order to identify the mitotic germ cells. However, little is known in fish about their cell surface antigen. In this study, we identified lymphocyte antigen 75 (Ly75/CD205) as a germ cell-specific cell surface marker by combination expressed sequence tag analysis of purified type A spermatogonia (A-SG) from immature testis, in silico prediction of membrane proteins, and expression studies. The ly75 transcripts were abundant in the testis and gills, and weak signals were detected in the head kidney and brain. In addition, ly75 mRNA was predominantly localized in the primordial germ cells of newly hatched embryos, A-SG in testis, oogonia, and chromatin nucleolus-stage oocytes in the ovary. In contrast, ly75 mRNA was not detected in spermatocytes, spermatids, spermatozoa, vitellogenic oocytes, or gonadal somatic cells from either males or females. The expression profile of Ly75 protein was similar to that of the mRNA. Furthermore, identification of various fish homologs of ly75 confirmed that their amino acid sequences are well conserved. Therefore, Ly75 may be appropriate for use as a versatile cell surface marker for mitotic germ cells in fish.
Biology of Reproduction 10/2010; 83(4):597-606. · 4.01 Impact Factor
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ABSTRACT: The production of xenogenic gametes from large-bodied, commercially important marine fish species in closely related smaller host fish species with short generation times may enable rapid and simple seed production of the target species. As a first step toward this goal, we assessed the suitability of chub mackerel, Scomber japonicus, as a small-bodied recipient species for xenogenic spermatogonial transplantation. Histological observation of the early gonadal development of chub mackerel larvae and transplantation of fluorescent-labeled spermatogonia from Nibe croaker, Nibea mitsukurii, revealed that 5.3-mm chub mackerel larvae were suitable recipients for successful transplantation. Intraperitoneally transplanted xenogenic spermatogonia efficiently colonized the gonads of these recipient larvae, and donor-derived Nibe croaker germ cells proliferated rapidly soon after colonization. Moreover, gonadal soma-derived growth factor (gsdf) mRNA, a gonadal somatic cell marker, was expressed in recipient-derived cells surrounding the incorporated donor-derived germ cells, suggesting that donor-derived germ cells had settled at an appropriate location in the recipient gonad. Our data show that xenogenic spermatogonial transplantation was successful in chub mackerel and that the somatic microenvironment of the chub mackerel gonad can support the colonization, survival, and proliferation of intraperitoneally transplanted xenogenic germ cells derived from a donor species of a different taxonomic family.
Biology of Reproduction 05/2010; 82(5):896-904. · 4.01 Impact Factor
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ABSTRACT: In a recently established system for intraperitoneal spermatogonial cell transplantation in salmonids, donor type A spermatogonia (type A SG) were microinjected into the peritoneal cavity of newly hatched larvae. Compared with salmonids, the larvae of marine teleosts are small and vulnerable to physiological and physical stresses, making it difficult to use them for cell manipulation. Herein, we developed type A SG cell transplantation in Nibe croaker (Nibea mitsukurii) by optimizing 1) the developmental stage of the donor testes used to prepare type A SG-enriched cell suspensions and 2) the timing and location of intraperitoneal cell transplantations to recipient larvae. Donor cells labeled with PKH26 fluorescent dye were transplanted into the peritoneal cavity of 3-, 4-, 5-, and 6-mm larvae using glass micropipettes. Consequently, 20.6% of the 4-mm larvae recipients survived for 3 wk, and 36.3% of the survivors had donor-derived cells in their gonads. The incorporated donor cells were identified as germ cells by germ cell-specific nuclear morphology and expression of a germ cell marker. In contrast, no donor type A SG were incorporated into the gonads of 6-mm recipient larvae. These data indicate that there is a distinct narrow window in the developmental stages of recipient larvae when exogenous type A SG can be incorporated into the gonads. The establishment of this system in pelagic egg-spawning marine teleosts would allow the creation of a new broodstock system in which a target species with a large body size and long generation time could be produced from related species with a small body size and short generation time.
Biology of Reproduction 08/2009; 81(6):1055-63. · 4.01 Impact Factor
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ABSTRACT: In this study, a Pacific bluefin tuna (Thunnus orientalis) homolog of the Drosophila vasa gene, BtVLG (bluefin tuna vasa-like gene), was cloned and characterized for use as a molecular marker for germ cells in this species. Analysis of the nucleotide sequence revealed that BtVLG comprises 2,394 bps with an open reading frame of 1,932 bps encoding 644 amino acids. The deduced amino acid sequence contained arginine-glycine or arginine-glycine-glycine motifs and eight conserved motifs belonging to the DEAD-box protein family. The BtVLG sequence showed high similarity to Drosophila vasa (69.1%), zebrafish vasa homolog (80.5%), and tilapia vasa homolog (91.2%). In adult tissues, the BtVLG transcripts were specifically detected in ovary and testis. In situ hybridization analysis showed that BtVLG messenger RNA (mRNA) was detected in oogonia and previtellogenic oocytes in the ovary. In the testis, while BtVLG mRNA was detected in spermatogonia, it was not detected in the primary and secondary spermatocytes, spermatids, spermatozoa or gonadal somatic cells. Consequently, consensus sequences, sequence similarity, and specific localization of BtVLG mRNA in the germ cells all suggest that BtVLG is the bluefin tuna vasa homolog of the Drosophila vasa gene. Further, BtVLG can be used as a molecular marker for bluefin tuna germ cells.
Fisheries Science 12/2008; 75(1):71-79. · 0.94 Impact Factor
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Tanpakushitsu kakusan koso. Protein, nucleic acid, enzyme 12/2007; 52(16 Suppl):2067-72.
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ABSTRACT: Many salmonids have become at risk of extinction. For teleosts whose eggs cannot be cryopreserved, developing techniques other than egg cryopreservation to save genetic resources is imperative. In this study, spermatogonia from rainbow trout were intraperitoneally transplanted into newly hatched sterile triploid masu salmon. Transplanted trout spermatogonia underwent spermatogenesis and oogenesis in male and female recipients, respectively. At 2 years after transplantation, triploid salmon recipients only produced trout sperm and eggs. With use of these salmon as parents, we successfully produced only donor-derived trout offspring. Thus, by transplanting cryopreserved spermatogonia into sterile xenogeneic recipients, we can generate individuals of a threatened species.
Science 10/2007; 317(5844):1517. · 31.20 Impact Factor
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ABSTRACT: An increasing number of wild fish species are in danger of extinction, often as a result of human activities. The cryopreservation of gametes and embryos has great potential for maintaining and restoring threatened species. The conservation of both paternal and maternal genetic information is essential. However, although this technique has been successfully applied to the spermatozoa of many fish species, reliable methods are lacking for the long-term preservation of fish eggs and embryos. Here, we describe a protocol for use with rainbow trout (Oncorhynchus mykiss) primordial germ cells (PGCs) and document the restoration of live fish from gametes derived from these cryopreserved progenitors. Genital ridges (GRs), which are embryonic tissues containing PGCs, were successfully cryopreserved in a medium containing 1.8 M ethylene glycol (EG). The thawed PGCs that were transplanted into the peritoneal cavities of allogenic trout hatchlings differentiated into mature spermatozoa and eggs in the recipient gonads. Furthermore, the fertilization of eggs derived from cryopreserved PGCs by cryopreserved spermatozoa resulted in the development of fertile F1 fish. This PGC cryopreservation technique represents a promising tool in efforts to save threatened fish species. Moreover, this approach has significant potential for maintaining domesticated fish strains carrying commercially valuable traits for aquaculture purposes.
Molecular Reproduction and Development 03/2007; 74(2):207-13. · 2.53 Impact Factor
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ABSTRACT: Members of the bone morphogenetic protein (BMP) family play diverse roles in multiple developmental processes. However, in the mouse, mutations in many BMPs, BMP receptors and signaling components result in early embryonic lethality making it difficult to analyze the role of these factors during organogenesis or tissue homeostasis in the adult. To bypass this early lethality, we used an organ culture system to study the role of BMPs during primordial germ cell (PGC) migration. PGCs are the embryonic precursors of the sperm and eggs. BMPs induce formation of primordial germ cells within the proximal epiblast of embryonic day 7.5 (E7.5) mouse embryos. PGCs then migrate via the gut to arrive at the developing gonads by E10.5. Addition of BMP4 or the BMP-antagonist Noggin to transverse slices dissected from E9.5 embryos elevated PGC numbers or reduced PGC numbers, respectively. Noggin treatment also slowed and randomized PGC movements, resulting in a failure of PGCs to colonize the urogenital ridges (UGRs). Based on p-Smad1/5/8 staining, migratory PGCs do not respond to endogenous BMPs. Instead, the somatic cells of the urogenital ridges exhibit elevated p-Smad1/5/8 staining revealing active BMP signaling within the UGRs. Noggin treatment abrogated p-Smad staining within the UGRs and blocked localized expression of Kitl, a cytokine known to regulate the survival and motility of PGCs and Id1, a transcription factor expressed within the UGRs. We propose that BMP signaling regulates PGC migration by controlling gene expression within the somatic cells along the migration route and within the genital ridges.
Mechanisms of Development 02/2007; 124(1):68-77. · 2.83 Impact Factor
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ABSTRACT: Germ-cell transplantation has many applications in biology and animal husbandry, including investigating the complex processes of germ-cell development and differentiation, producing transgenic animals by genetically modifying germline cells, and creating broodstock systems in which a target species can be produced from a surrogate parent. The germ-cell transplantation technique was initially established in chickens using primordial germ cells (PGCs), and was subsequently extended to mice using spermatogonial stem cells. Recently, we developed the first germ-cell transplantation system in lower vertebrates using fish PGCs and spermatogonia. During mammalian germ-cell transplantation, donor spermatogonial stem cells are introduced into the seminiferous tubules of the recipient testes. By contrast, in the fish germ-cell transplantation system, donor cells are microinjected into the peritoneal cavities of newly hatched embryos; this allows the donor germ cells to migrate towards, and subsequently colonize, the recipient genital ridges. The recipient embryos have immature immune systems, so the donor germ cells can survive and even differentiate into mature gametes in their allogeneic gonads, ultimately leading to the production of normal offspring. In addition, implanted spermatogonia can successfully differentiate into sperm and eggs, respectively, in male and female recipients. The results of transplantation studies in fish are improving our understanding of the development of germ-cell systems during vertebrate evolution.
Journal of Reproduction and Development 01/2007; 52(6):685-93. · 1.46 Impact Factor
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ABSTRACT: Neurons that synthesize and release GnRH are essential for the central regulation of reproduction. Evidence suggests that forebrain GnRH neurons originate in the olfactory placode and migrate to their final destinations, although this is still a matter of controversy. X-linked Kallmann syndrome (X-KS), characterized by failed gonadal function secondary to deficient gonadotropin secretion, is caused by a mutation in KAL1, which is suggested to regulate the migration of forebrain GnRH neurons. Because rodents lack Kal1 in their genome and have GnRH neurons scattered throughout their forebrain, the development of forebrain GnRH neurons and the pathogenesis of X-KS have been difficult to study. In the present study, we generated transgenic medaka that expressed green fluorescent protein under the control of the gnrh1 and gnrh3 promoters for analyzing forebrain GnRH neuronal development. Our data revealed the presence of the following four gnrh1 neuronal populations: an olfactory region-derived ventral preoptic population, a dorsal preoptic population that migrates from the dorsal telencephalon, a medial ventral telencephalic population that migrates from the anterior telencephalon, and a nonmigratory ventral hypothalamic population. We found that all forebrain gnrh3 neurons, extending from the terminal nerve ganglion to the anterior mesencephalon, arise from the olfactory region and that trigeminal ganglion neurons express gnrh3. Maternal gnrh3 expression was also observed in oocytes and early embryos. We subsequently identified a KAL1 ortholog and its paralogous form in the medaka. Consistent with the X-KS phenotype, antisense knockdown of the medaka KAL1 ortholog resulted in the disruption of forebrain GnRH neuronal migration. Thus, these transgenic medaka provide a useful model system for studying GnRH neuronal development and disorders of GnRH deficiency.
Endocrinology 04/2006; 147(3):1076-84. · 4.46 Impact Factor
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ABSTRACT: Understanding the mechanisms that regulate germ-cell development is crucial to reproductive medicine and animal production. Animal gametes originally derive from sexually undifferentiated primordial germ cells (PGCs), which develop into mitotic germ cells (oogonia or spermatogonia) before proceeding to meiosis [Wylie, C. (1999) Cell 96, 165-174]. Spermatogonia are thought to include a population of cells with stem cell activity, which proliferate throughout the lifespan of male animals and produce spermatozoa [Zhao, G. Q. & Garbers, D. L. (2002) Dev. Cell 2, 537-547]. However, the functional differences between PGCs and spermatogonial stem cells are poorly understood. Here we show that transplanted adult testicular germ cells can colonize sexually undifferentiated embryonic gonads and resume gametogenesis. Testicular germ cells containing spermatogonial stem cells isolated from adult male rainbow trout (Oncorhynchus mykiss) were transplanted into the peritoneal cavity of newly hatched embryos of both sexes, and the behavior of the donor cells was observed. The testicular germ cells differentiated into spermatozoa in male recipients and fully functional eggs in female recipients. Furthermore, the donor-derived spermatozoa and eggs obtained from the recipient fish were able to produce normal offspring. These findings indicate that fish testicular germ cells, probably spermatogonial stem cells, possess a high level of developmental plasticity and sexual bipotency, even after the animal reaches maturity. Furthermore, our results suggest that spermatogonial stem cells are at least partly functionally similar to PGCs.
Proceedings of the National Academy of Sciences 02/2006; 103(8):2725-9. · 9.68 Impact Factor
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ABSTRACT: Fibroblast growth factor (FGF) signaling is thought to play a role in germ cell behavior. FGF2 has been reported to be a mitogen for primordial germ cells in vitro, whilst combinations of FGF2, steel factor and LIF cause cultured germ cells to transform into permanent lines of pluripotent cells resembling ES cells. However, the actual function of FGF signaling on the migrating germ cells in vivo is unknown. We show, by RT-PCR analysis of cDNA from purified E10.5 germ cells, that germ cells express two FGF receptors: Fgfr1-IIIc and Fgfr2-IIIb. Second, we show that FGF-mediated activation of the MAP kinase pathway occurs in germ cells during their migration, and thus they are potentially direct targets of FGF signaling. Third, we use cultured embryo slices in simple gain-of-function experiments, using FGF ligands, to show that FGF2, a ligand for FGFR1-IIIc, affects motility, whereas FGF7, a ligand for FGFR2-IIIb, affects germ cell numbers. Loss of function, using a specific inhibitor of FGF signaling, causes increased apoptosis and inhibition of cell shape change in the migrating germ cells. Lastly, we confirm in vivo the effects seen in slice cultures in vitro, by examining germ cell positions and numbers in embryos carrying a loss-of-function allele of FGFR2-IIIb. In FGFR2-IIIb(-/-) embryos, germ cell migration is unaffected, but the numbers of germ cells are significantly reduced. These data show that a major role of FGF signaling through FGFR2-IIIb is to control germ cell numbers. The data do not discriminate between direct and indirect effects of FGF signaling on germ cells, and both may be involved.
Development 01/2006; 132(24):5399-409. · 6.60 Impact Factor
