An engineered 3D blood-testis barrier model for the assessment of reproductive toxicity potential

Legendre, Audrey; Froment, Pascal; Desmots, Sophie; Lecomte, Anthony; Habert, René; Lemazurier, Emmanuel

doi:10.1016/j.biomaterials.2010.02.029

Cited by 52 publications

(34 citation statements)

References 79 publications

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“…The organ culture of rat testicular tissue was performed for up to 52 days, to ensure that the whole process of spermatogenesis, which in vivo takes 52–53 days in rats, starting from type A spermatogonia, present at the age of 5 d pp in rats (Clermont and Harvey, 1965; Legendre et al ., 2010), could be completed within the culture period. The culture medium, MEMα with 10% (v/v) KSR, was used as it has been shown earlier to support male germ cell differentiation in vitro in mice (Sato et al ., 2011).…”

Section: Discussionmentioning

confidence: 99%

In vitrodifferentiation of rat spermatogonia into round spermatids in tissue culture

et al. 2016

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STUDY QUESTIONDo the organ culture conditions, previously defined for in vitro murine male germ cell differentiation, also result in differentiation of rat spermatogonia into post-meiotic germ cells exhibiting specific markers for haploid germ cells?SUMMARY ANSWERWe demonstrated the differentiation of rat spermatogonia into post-meiotic cells in vitro, with emphasis on exhibiting, protein markers described for round spermatids.WHAT IS KNOWN ALREADYFull spermatogenesis in vitro from immature germ cells using an organ culture technique in mice was first reported 5 years ago. However, no studies reporting the differentiation of rat spermatogonia into post-meiotic germ cells exhibiting the characteristic protein expression profile or into functional sperm have been reported.STUDY DESIGN, SAMPLES/MATERIALS, METHODSOrgan culture of testicular fragments of 5 days postpartum (dpp) neonatal rats was performed for up to 52 days. Evaluation of microscopic morphology, testosterone levels, mRNA and protein expression as measured by RT-qPCR and immunostaining were conducted to monitor germ cell differentiation in vitro. Potential effects of melatonin, Glutamax® medium, retinoic acid and the presence of epidydimal fat tissue on the spermatogenic process were evaluated. A minimum of three biological replicates were performed for all experiments presented in this study. One-way ANOVA, ANOVA on ranks and student's t-test were applied to perform the statistical analysis.MAIN RESULTS AND THE ROLE OF CHANCEMale germ cells, present in testicular tissue pieces grown from 5 dpp rats, exhibited positive protein expression for Acrosin and Crem (cAMP (cyclic adenosine mono phosphate) response element modulator) after 52 days of culture in vitro. Intra-testicular testosterone production could be observed after 3 days of culture, while when epididymal fat tissue was added, spontaneous contractility of cultured seminiferous tubules could be observed after 21 days. However, no supportive effect of the supplementation with any factor or the co-culturing with epididymal fat tissue on germ cell differentiation in vitro or testosterone production was observed.LIMITATIONS, REASONS FOR CAUTIONThe human testis is very different in physiology from the rat testis, further investigations are still needed to optimize the organ culture system for future use in humans.WIDER IMPLICATIONS OF THE FINDINGSThe successful differentiation of undifferentiated spermatogonia using the testis explant culture system might be employed in future to produce sperm from human spermatogonia as a clinical tool for fertility preservation in boys and men suffering infertility.LARGE SCALE DATANone.STUDY FUNDING AND COMPETING INTEREST(S)This work was supported financially by the Frimurare Barnhuset in Stockholm, the Paediatric Research Foundation, Jeanssons Foundation, Sällskåpet Barnåvard in Stockholm, Swedish Research Council/Academy of Finland, Emil and Wera Cornells Foundation, Samariten Foundation, the Swedish Childhood Cancer Foundation as well as through the regional agre...

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Section: Discussionmentioning

confidence: 99%

In vitrodifferentiation of rat spermatogonia into round spermatids in tissue culture

et al. 2016

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“…Seminiferous tubules were prepared as described previously (Guibert et al 2013) and seeded in static 24-well cell culture polyethylene terephthalate inserts (pore 0.4 µm, WWR, Fontenay sous Bois, France) as described previously (Djakiew & Dym 1988, Staub et al 2000, Legendre et al 2010. This was carried out in order to preserve germ cells on polarised Sertoli cells in a dynamic culture using the Quasi-Vivo system (Kirkstall Ltd, Rotherham, UK).…”

Section: Culture Of Chicken Seminiferous Tubulesmentioning

confidence: 99%

The insulin sensitiser metformin regulates chicken Sertoli and germ cell populations

Faure¹,

Guibert²,

Alves³

et al. 2016

Reproduction

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Metformin, an insulin sensitiser from the biguanide family of molecules, is used for the treatment of insulin resistance in type 2 diabetes individuals. It increases peripheral glucose uptake and may reduce food intake. Based on the tight link between metabolism and fertility, we investigated the role of metformin on testicular function using in vitro culture of Sertoli cells and seminiferous tubules, complemented by in vivo data obtained following metformin administration to prepubertal chickens. In vitro, metformin treatment reduced Sertoli cell proliferation without inducing apoptosis and morphological changes. The metabolism of Sertoli cells was affected because lactate secretion by Sertoli cells increased approximately twofold and intracellular free ATP was negatively impacted. Two important pathways regulating proliferation and metabolism in Sertoli cells were assayed. Metformin exposure was not associated with an increased phosphorylation of AKT or ERK. There was a 90% reduction in the proportion of proliferating germ cells after a 96-h exposure of seminiferous tubule cultures to metformin. In vivo, 6-week-old chickens treated with metformin for 3 weeks exhibited reduced testicular weight and a 50% decrease in testosterone levels. The expression of a marker of undifferentiated germ cells was unchanged in contrast to the decrease in expression of 'protamine', a marker of differentiated germ cells. In conclusion, these results suggest that metformin affects the testicular energy content and the proliferative ability of Sertoli and germ cells.

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“…Likewise, there were other studies using 3D cell culture system to gain more insights into the interactions among germ cells, somatic cells and various extracellular matrices for the in vitro spermatogenesis (Hue et al 1998, Tanaka et al 2003, Stukenborg et al 2009, Abu Elhija et al 2012. The 3D blood-testis barrier model was also used for rat germ cell differentiation with production of haploid cells (Legendre et al 2010). A novel organ culture condition has been shown to support the whole spermatogenesis of neonatal mouse testicular issues and cultured with serum-free medium (Sato et al 2011a).…”

Section: Male Differentiated Germ Cells Derivation From Sscsmentioning

confidence: 99%

Generation of male differentiated germ cells from various types of stem cells

Hou

Yang

et al. 2014

Reproduction

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Infertility is a major and largely incurable disease caused by disruption and loss of germ cells. It affects 10-15% of couples, and male factor accounts for half of the cases. To obtain human male germ cells 'especially functional spermatids' is essential for treating male infertility. Currently, much progress has been made on generating male germ cells, including spermatogonia, spermatocytes, and spermatids, from various types of stem cells. These germ cells can also be used in investigation of the pathology of male infertility. In this review, we focused on advances on obtaining male differentiated germ cells from different kinds of stem cells, with an emphasis on the embryonic stem (ES) cells, the induced pluripotent stem (iPS) cells, and spermatogonial stem cells (SSCs). We illustrated the generation of male differentiated germ cells from ES cells, iPS cells and SSCs, and we summarized the phenotype for these stem cells, spermatocytes and spermatids. Moreover, we address the differentiation potentials of ES cells, iPS cells and SSCs. We also highlight the advantages, disadvantages and concerns on derivation of the differentiated male germ cells from several types of stem cells. The ability of generating mature and functional male gametes from stem cells could enable us to understand the precise etiology of male infertility and offer an invaluable source of autologous male gametes for treating male infertility of azoospermia patients.

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An engineered 3D blood-testis barrier model for the assessment of reproductive toxicity potential

Cited by 52 publications

References 79 publications

In vitrodifferentiation of rat spermatogonia into round spermatids in tissue culture

In vitrodifferentiation of rat spermatogonia into round spermatids in tissue culture

The insulin sensitiser metformin regulates chicken Sertoli and germ cell populations

Generation of male differentiated germ cells from various types of stem cells

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