scholarly journals Germ cell development in equine testis tissue xenografted into mice

Reproduction ◽  
2006 ◽  
Vol 131 (6) ◽  
pp. 1091-1098 ◽  
Author(s):  
R Rathi ◽  
A Honaramooz ◽  
W Zeng ◽  
R Turner ◽  
I Dobrinski
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Meiotic Division ◽  
Seminal Vesicles ◽  
Post Transplantation ◽  
Germ Cell Development ◽  
Immunodeficient Mice ◽  
Low Efficiency ◽  
Underlying Mechanisms ◽  
Testis Tissue

Grafting of testis tissue from immature animals to immunodeficient mice results in complete spermatogenesis, albeit with varying efficiency in different species. The objectives of this study were to investigate if grafting of horse testis tissue would result in spermatogenesis, and to assess the effect of exogenous gonadotropins on xenograft development. Small fragments of testis tissue from 7 colts (2 week to 4 years of age) were grafted under the back skin of castrated male immunodeficient mice. For 2 donor animals, half of the mice were treated with gonadotropins. Xenografts were analyzed at 4 and 8 months post-transplantation. Spermatogenic differentiation following grafting ranged from no differentiation to progression through meiosis with appearance of haploid cells. Administration of exogenous gonadotropins appeared to support post-meiotic differentiation. For more mature donor testis samples where spermatogenesis had progressed into or through meiosis, after grafting an initial loss of differentiated germ cells was observed followed by a resurgence of spermatogenesis. However, if haploid cells had been present prior to grafting, spermatogenesis did not progress beyond meiotic division. In all host mice with spermatogenic differentiation in grafts, increased weight of the seminal vesicles compared to castrated mice showed that xenografts were releasing testosterone. These results indicate that horse spermatogenesis occurs in a mouse host albeit with low efficiency. In most cases, spermatogenesis arrested at meiosis. The underlying mechanisms of this spermatogenic arrest require further investigation.

scholarly journals 193 TESTIS TISSUE XENOGRAFTING TO PRESERVE GERM CELLS FROM A CLONED BANTENG CALF

2005 ◽  
Vol 17 (2) ◽  
pp. 247 ◽  
Author(s):  
A. Honaramooz ◽  
W. Zeng ◽  
R. Rathi ◽  
J. Koster ◽  
O. Ryder ◽  
...  
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Seminal Vesicles ◽  
Male Germ Cells ◽  
Newborn Animal ◽  
Immunodeficient Mice ◽  
Germ Cell Differentiation ◽  
Pachytene Spermatocytes ◽  
The Right ◽  
Testis Tissue

In April 2003, two banteng (Bos javonicus) calves were born after heterologous nuclear transfer of donor cells from a genetically valuable individual frozen in 1978. One of the cloned banteng calves died at one week of age. The calf was found to have one scrotal and one abdominally cryptorchid testis. In an attempt to preserve male germ cells from this valuable animal, parts of each testis were shipped on ice to the University of Pennsylvania for xenografting. Grafting of testis tissue from immature domestic animals and monkeys under the back skin of immunodeficient mice can result in complete spermatogenesis, albeit with different levels of efficiency in different species. The objective of this experiment was to investigate if grafting of immature banteng testis tissue would result in spermatogenesis in a mouse host. Small fragments of tissue (about 1 mm, 3 each) from both testes were grafted under the back skin (4 pieces of scrotal testis on the right side and 4 pieces of retained testis on the left side) of 6 castrated male immunodeficient mice. Histological examination of the testis xenografts was performed 3, 6, 9, 12, and 15 months after transplantation. Weight of the seminal vesicles in the host mouse was recorded as an indicator of bioactive testosterone produced by the xenografts. At the time of grafting, both testes contained seminiferous cords with immature Sertoli cells and gonocytes. At 3, 6, and 9 months after grafting, pachytene spermatocytes were present in the xenografts of the scrotal testis whereas no germ cell differentiation was observed in grafts from the retained testis. However, spermatogenesis in grafts of the scrotal testis did not proceed further through meiosis in grafts analyzed at 12 and 15 months after grafting, with pachytene spermatocytes still the most advanced germ cell type present in grafts recovered 15 months after grafting. The weight of the seminal vesicles in the castrated host mice was restored to pre-castration values showing that xenografts were releasing bioactive testosterone. These results indicate that banteng spermatogenesis was initiated in the mouse host but became arrested at meiosis as observed previously in xenografts of immature bovine or equine testis. Therefore, haploid germ cells could not be recovered. This represents the first example of trying to preserve fertility from a rare, valuable newborn animal by testis tissue xenografting. While xenografting presents a previously unavailable option for preservation of male germ cells from immature individuals, the efficiency of sperm production in testis xenografts appears to be variable and has to be determined empirically for different donor species. This work was supported by USDA 03-35203-13486.

scholarly journals 268 GERM CELL DEVELOPMENT IN EQUINE TESTIS TISSUE XENOGRAFTED INTO MICE

2005 ◽  
Vol 17 (2) ◽  
pp. 283 ◽  
Author(s):  
R. Turner ◽  
R. Rathi ◽  
A. Honaramooz ◽  
W. Zeng ◽  
I. Dobrinski
Keyword(s):  
Chorionic Gonadotropin ◽  
Germ Cells ◽  
Seminal Vesicles ◽  
Post Transplantation ◽  
Immunodeficient Mice ◽  
Donor Tissue ◽  
Old Donor ◽  
Pachytene Spermatocytes ◽  
Donor Species ◽  
Testis Tissue

Grafting of testis tissue from immature animals under the back skin of immunodeficient mice results in complete spermatogenesis, albeit with different levels of efficiency in different species. While spermatogenesis develops comparably to that in the donor species in xenografts from pigs, sheep and goats, spermatogenic differentiation is less efficient in testis tissue from cats and bulls. Testicular maturation was significantly accelerated in rhesus monkey testis grafts whereas timing was similar to that in the donor species in cats and bulls. The objective of this study was to investigate if grafting of immature horse testis tissue would result in spermatogenesis in a mouse host. Small fragments of testis tissue (about 1 mm3) from four sexually immature colts (2-week-old Standardbred, 5- and 8-month-old ponies, 10-month-old Warmblood) were grafted under the back skin of castrated male immunodeficient mice (n = 5, 5, 10 and 5 recipient mice, respectively). Histological examination of the testis xenografts was performed between 14 and 50 week post-transplantation. Weight of the seminal vesicles in the host mouse was recorded as an indicator of bioactive testosterone produced by the xenografts. At the time of grafting, the seminiferous cords of the donor testis tissue form 2-week-, 5-month- and 8-month-old colts contained only immature Sertoli cells and gonocytes. No spermatogenic differentiation occurred in xenografts from the 2-week-old colt and testosterone production was minimal. Pachytene spermatocytes were observed in testis grafts from the 5- and 8-month-old donors from 14 weeks onward. Spermatogenesis did not proceed through meiosis in grafts from the 5-month-old donor. Recipient mice carrying xenografts from the 8-month-old donor received exogenous gonadotropins (equine chorionic gonadotropin and human chorionic gonadotropin, 10 I.U./day for 2 months, beginning 14 weeks after grafting) and condensing spermatids were observed by 35 weeks after grafting. In donor tissue from the 10-month-old colt, pachytene spermatocytes were present in about 50% of tubules at the time of grafting. After 14 weeks, xenografts showed fewer differentiated germ cells than the donor tissue. However, at 35 weeks after grafting, condensing spermatids were observed, indicating that differentiated germ cells were initially lost and spermatogenesis was subsequently reinitiated. In all castrated host mice where spermatogenic differentiation occurred, the weight of the seminal vesicles was restored to pre-castration values showing that xenografts were releasing bioactive testosterone. These results indicate that horse spermatogenesis can occur in a mouse host albeit with low efficiency. Testicular maturation was not accelerated. In most cases, spermatogenesis appeared to become arrested at meiosis. The underlying mechanisms of this spermatogenic arrest require further investigation. Although equine testis xenografts produced testosterone, supplementation of exogenous gonadotropins might support post-meiotic differentiation. This work was supported by USDA 03-35203-13486.

scholarly journals Germ cell fate and seminiferous tubule development in bovine testis xenografts

Reproduction ◽  
2005 ◽  
Vol 130 (6) ◽  
pp. 923-929 ◽  
Author(s):  
Rahul Rathi ◽  
Ali Honaramooz ◽  
Wenxian Zeng ◽  
Stefan Schlatt ◽  
Ina Dobrinski
Keyword(s):  
Germ Cell ◽  
Cell Fate ◽  
Germ Cells ◽  
Cell Number ◽  
Sperm Production ◽  
Continuous Increase ◽  
Control Tissue ◽  
Low Efficiency ◽  
Pachytene Spermatocytes ◽  
Testis Tissue

Spermatogenesis can occur in testis tissue from immature bulls ectopically grafted into mouse hosts; however, efficiency of sperm production is lower than in other donor species. To elucidate a possible mechanism for the impaired spermatogenesis in bovine testis xenografts, germ cell fate and xenograft development were investigated at different time points and compared with testis tissue from age-matched calves as controls. Histologically, an initial decrease in germ cell number was noticed in xenografts recovered up to 2 months post-grafting without an increase in germ cell apoptosis. From 2 months onward, the number of germ cells increased. In contrast, a continuous increase in germ cell number was seen in control tissue. Pachytene spermatocytes were observed in some grafts before 4 months, whereas in the control tissue they were not present until 5 months of age. Beyond 4 months post-grafting spermatogenesis appeared to be arrested at the pachytene spermatocyte stage in most grafts. Elongated spermatids were observed between 6 and 8 months post-grafting, similar to the controls, albeit in much lower numbers. Lumen formation started earlier in grafts compared with controls and by 6 months post-grafting tubules with extensively dilated lumen were observed. A donor effect on efficiency of spermatogenesis was also observed. These results indicate that the low efficiency of sperm production in bovine xenografts is due to an initial deficit of germ cells and impaired meiotic and post-meiotic differentiation. The characterization of spermatogenic efficiency will provide the basis to understand the control of spermatogenesis in testis grafts.

scholarly journals 267 TESTIS TISSUE XENOGRAFTING AS A BIOASSAY FOR GERM CELL DEVELOPMENTAL POTENTIAL IN EQUINE CRYPTORCHID TESTES

2005 ◽  
Vol 17 (2) ◽  
pp. 283 ◽  
Author(s):  
R. Rathi ◽  
A. Honaramooz ◽  
W. Zeng ◽  
R. Turner ◽  
I. Dobrinski
Keyword(s):  
Body Temperature ◽  
Germ Cells ◽  
Core Body Temperature ◽  
Domestic Animals ◽  
Seminal Vesicles ◽  
Developmental Potential ◽  
Immunodeficient Mice ◽  
Core Body ◽  
Testis Tissue

In domestic animals, spermatogenic differentiation is blocked in abdominally retained testes exposed to core body temperature. It is not known if undifferentiated germ cells are retained in long-term cryptorchid equine testes, nor is it known whether any surviving germ cells retain their ability to progress through spermatogenesis. If functional germ cells do persist in equine abdominal testes, then the possibility exists that offspring could be derived even from bilaterally cryptorchid individuals. Previously, we reported an in vivo model where completion of spermatogenesis with production of spermatozoa capable of fertilization occurred in fragments of testicular tissue from immature mice, domestic animals, and monkeys grafted under the skin of immunodeficient mice. Therefore, spermatogenic development in testis tissue xenografts can serve as an in vivo assay system for the developmental potential of germ cells. The objective of this study was to investigate if cryptorchid horse testes that had been exposed to core body temperature for 1–3 years had retained developmentally competent germ cells. Small fragments of abdominally cryptorchid testis tissue (about 1 mm3) from three donor horses (1-, 2-, and 3-year-old Quarterhorse) were grafted under the back skin of castrated male immunodeficient mice (n = 8, 6, and 3 recipient mice, respectively). At the time of grafting, donor tissue did not contain differentiated germ cells. Histological examination of the testis xenografts was performed between 5 and 45 weeks post-transplantation. Weight of the seminal vesicles in the host mouse was recorded as an indicator of bioactive testosterone produced by the xenografts. By 28 weeks after grafting, pachytene spermatocytes were observed in xenografts from all cryptorchid donor testes. While haploid gametes would be expected to be present in xenografted testis tissue from descended equine testes by 35 weeks after grafting, spermatogenesis did not progress through meiosis in the cryptorchid grafts. In all recipient animals where spermatogenic differentiation occurred, the weight of the seminal vesicles in the castrated host mice was restored to pre-castration values, indicating that xenografts were capable of releasing biologically active testosterone. These results indicate that even after 3 years of exposure to core body temperature, equine cryptorchid testes contain germ cells capable of differentiation. It remains to be investigated if supplementation of exogenous gonadotropins might support post-meiotic differentiation of germ cells in cryptorchid equine testes xenografts. This work was supported by USDA 03-35203-13486.

scholarly journals Establishment of Etv5 gene knockout mice as a recipient model for spermatogonial stem cell transplantation

2020 ◽  
Author(s):  
Xianyu Zhang ◽  
Xin Zhao ◽  
Guoling Li ◽  
Mao Zhang ◽  
Pingping Xing ◽  
...  
Keyword(s):  
Germ Cell ◽  
Cell Transplantation ◽  
Germ Cells ◽  
Fluorescent Protein ◽  
Gene Knockout ◽  
Seminiferous Tubules ◽  
Post Transplantation ◽  
Mature Adult ◽  
Testicular Weight ◽  
Low Efficiency

AbstractTransplantation of spermatogonial stem cells (SSCs) is an alternative reproductive method to achieve conservation and production of elite animals in livestock production. Creating a recipient animal without endogenous germ cells is important for effective SSC transplantation. However, natural mutants with depletion of SSCs are difficult to obtain, and drug ablation of endogenous germ cells is arduous to perform for practical use. In this study, we used mouse models to study the preparation of recipients with congenital germ cell ablation. We knocked out (KO) Ets-variant gene 5 (Etv5) in mice using the CRISPR/Cas9 system. The testicular weight of Etv5-/- mice was significantly lower than that of wild-type (WT) mice. The germ cell layer of the seminiferous tubules gradually receded with age in Etv5-/- mice. At 12 weeks of age, the tubules of Etv5-/- mice lacked germ cells (Sertoli cell-only syndrome), and sperm were completely absent in the epididymis. We subsequently transplanted allogeneic SSCs with enhanced green fluorescent protein (EGFP) into 3-(immature) or 7-week-old (mature) Etv5-/- mice. Restoration of germ cell layers in the seminiferous tubules and spermatogenesis was observed in all immature testes but not in mature adult testes at 2 months post-transplantation. The presence of heterologous genes Etv5 and EGFP in recipient testicular tissue and epididymal sperm by PCR indicated that sperm originated from the transplanted donor cells. Our study demonstrates that, although Etv5-/- mice could accommodate and support foreign germ cell transplantation, this process occurs in a quite low efficiency to support a full spermatogenesis of transplanted SSCs. However, using Etv5-/- mice as a recipient model for SSC transplantation is feasible, and still needs further investigation to establish an optimized transplantation process.

scholarly journals Loss of Kallmann syndrome-associated gene WDR11 disrupts primordial germ cell development by affecting canonical and non-canonical Hedgehog signalling

2020 ◽  
Author(s):  
Jiyoung Lee ◽  
Yeonjoo Kim ◽  
Paris Ataliotis ◽  
Hyung-Goo Kim ◽  
Dae-Won Kim ◽  
...  
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Primary Cilia ◽  
Primordial Germ Cell ◽  
Kallmann Syndrome ◽  
Loss Of Function ◽  
Downstream Effectors ◽  
Cell Components ◽  
Underlying Mechanisms ◽  
And Migration

ABSTRACTMutations of WDR11 are associated with Kallmann syndrome (KS) and congenital hypogonadotrophic hypogonadism (CHH), typically caused by defective functions of gonadotrophin-releasing hormone (GnRH) neurones in the brain. We previously reported that Wdr11 knockout mice show profound infertility with significantly fewer germ cells present in the gonads. To understand the underlying mechanisms mediated by WDR11 in these processes, we investigated the effects of Wdr11 deletion on primordial germ cell (PGC) development. Using live-tracking of PGCs and primary co-cultures of genital ridges (GR), we demonstrated that Wdr11-deficient embryos contained reduced numbers of PGCs which had delayed migration due to significantly decreased proliferation and motility. We found primary cilia-dependent canonical Hedgehog (Hh) signalling was required for proliferation of the somatic mesenchymal cells of GR, while primary cilia-independent non-canonical Hh signalling mediated by Ptch2/Gas1 and downstream effectors Src and Creb was required for PGC proliferation and migration, which was disrupted by the loss of function mutations of WDR11. Therefore, canonical and non-canonical Hh signalling are differentially involved in the development of somatic and germ cell components of the gonads, and WDR11 is required for both of these pathways operating in parallel in GR and PGCs, respectively, during normal PGC development. Our study provides a mechanistic link between the development of GnRH neurones and germ cells mediated by WDR11, which may underlie some cases of KS/CHH and ciliopathies.

scholarly journals Disruption of Tcfap2c in Primordial Germ Cells using Prdm1-Cre Prevents Germ Cell Development

2008 ◽  
Vol 78 (Suppl_1) ◽  
pp. 64-64
Author(s):  
Jillian Guttormsen ◽  
Gerrit J. Bouma ◽  
Frances Bhushan ◽  
Trevor Williams ◽  
Quinton A. Winger
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Primordial Germ Cells ◽  
Cell Development ◽  
Germ Cell Development

scholarly journals hnRNPH1 recruits PTBP2 and SRSF3 to cooperatively modulate alternative pre-mRNA splicing in germ cells and is essential for spermatogenesis and oogenesis

2021 ◽  
Author(s):  
Shuiqiao Yuan ◽  
Shenglei Feng ◽  
Jinmei Li ◽  
Hui Wen ◽  
Kuan Liu ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Germ Cell ◽  
Germ Cells ◽  
Rna Binding ◽  
Cell Development ◽  
Mrna Splicing ◽  
Conditional Knockout ◽  
Cell Junction ◽  
Germ Cell Development ◽  
Splicing Regulators

Abstract Coordinated regulation of alternative pre-mRNA splicing is essential for germ cell development. However, the molecular mechanism underlying that control alternative mRNA expression during germ cell development remains poorly understood. Herein, we showed that hnRNPH1, an RNA-binding protein, is highly expressed in the reproductive system and localized in the chromosomes of meiotic cells but excluded from the XY body in pachytene spermatocytes and recruits the splicing regulators PTBP2 and SRSF3 and cooperatively regulates the alternative splicing of the critical genes that are required for spermatogenesis. Conditional knockout Hnrnph1 in spermatogenic cells caused many abnormal splicing events that affect genes related to meiosis and communication between germ cells and Sertoli cells, characterized by asynapsis of chromosomes and impairment of germ-Sertoli communications, ultimately leading to male sterility. We further showed that hnRNPH1 could directly bind to SPO11 and recruit the splicing regulators PTBP2 and SRSF3 to regulate the alternative splicing of the target genes cooperatively. Strikingly, Hnrnph1 germline-specific mutant female mice were also infertile, and Hnrnph1-deficient oocytes exhibited a similar defective synapsis and cell-cell junction as shown in Hnrnph1-deficient male germ cells. Collectively, our data reveal an essential role for hnRNPH1 in regulating pre-mRNA splicing during spermatogenesis and oogenesis and support a molecular model whereby hnRNPH1 governs a network of alternative splicing events in germ cells via recruiting PTBP2 and SRSF3.

Isolation of the murine cyclin B2 cDNA and characterization of the lineage and temporal specificity of expression of the B1 and B2 cyclins during oogenesis, spermatogenesis and early embryogenesis

Development ◽  
1993 ◽  
Vol 118 (1) ◽  
pp. 229-240 ◽  
Author(s):  
D.L. Chapman ◽  
D.J. Wolgemuth
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Northern Blot ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Development ◽  
Early Embryogenesis ◽  
Northern Blot Hybridization ◽  
Germ Cell Development

A cDNA encoding the murine cyclin B2 (cycB2) was isolated from an adult mouse testis cDNA library as part of studies designed to identify cyclins involved in murine germ cell development. This cycB2 cDNA was then used to examine the pattern of cycB2 expression during male and female germ cell development and in early embryogenesis, and to compare this expression with the previously characterized expression of cycB1. A single 1.7 kb cycB2 transcript was detected by northern blot hybridization analysis of total RNA isolated from midgestation embryos and various adult tissues. Northern blot and in situ hybridization analyses revealed that cycB2 expression in the testis was most abundant in the germ cells, specifically in pachytene spermatocytes. This is in contrast to the highest levels of expression of cycB1 being present in early spermatids. In situ analysis of the ovary revealed cycB2 transcripts in both germ cells and somatic cells, specifically in the oocytes and granulosa cells of growing and mature follicles. The pattern of cycB1 and cycB2 expression in ovulated and fertilized eggs was also examined. While the steady state level of cycB1 and cycB2 signal remained constant in oocytes and ovulated eggs, signal of both appeared to decrease following fertilization. In addition, both cycB1 and cycB2 transcripts were detected in the cells of the inner cell mass and the trophectoderm of the blastocyst. These results demonstrate that there are lineage- and developmental-specific differences in the pattern of the B cyclins in mammalian germ cells, in contrast to the co-expression of B cyclins in the early conceptus.

scholarly journals Vascular endothelial growth factor regulates germ cell survival during establishment of spermatogenesis in the bovine testis

Reproduction ◽  
2009 ◽  
Vol 138 (4) ◽  
pp. 667-677 ◽  
Author(s):  
Kyle C Caires ◽  
Jeanene de Avila ◽  
Derek J McLean
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Growth Factor ◽  
Germ Cell ◽  
Cell Survival ◽  
Germ Cells ◽  
Pcr Analysis ◽  
Vascular Endothelial ◽  
Endothelial Growth Factor ◽  
Testis Tissue

Vascular endothelial growth factor-A (VEGFA) is a hypoxia-inducible peptide essential for angiogenesis and targets nonvascular cells in a variety of tissues and cell types. The objective of the current study was to determine the function of VEGF during testis development in bulls. We used an explant tissue culture and treatment approach to test the hypothesis that VEGFA-164 could regulate the biological activity of bovine germ cells. We demonstrate that VEGFA, KDR, and FLT1 proteins are expressed in germ and somatic cells in the bovine testis. Treatment of bovine testis tissue with VEGFA in vitro resulted in significantly more germ cells following 5 days of culture when compared with controls. Quantitative real-time RT-PCR analysis determined that VEGF treatment stimulated an intracellular response that prevents germ cell death in bovine testis tissue explants, as indicated by increased expression of BCL2 relative to BAX and decreased expression of BNIP3 at 3, 6, and 24 h during culture. Blocking VEGF activity in vitro using antisera against KDR and VEGF significantly reduced the number of germ cells in VEGF-treated testis tissue to control levels at 120 h. Testis grafting provided in vivo evidence that bovine testis tissue treated with VEGFA for 5 days in culture contained significantly more differentiating germ cells compared with controls. These findings support the conclusion that VEGF supports germ cell survival and sperm production in bulls.

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