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1.
Hum Reprod ; 38(3): 359-370, 2023 03 01.
Artículo en Inglés | MEDLINE | ID: mdl-36708005

RESUMEN

STUDY QUESTION: What is the impact of cancer or hematological disorders on germ cells in pediatric male patients? SUMMARY ANSWER: Spermatogonial quantity is reduced in testes of prepubertal boys diagnosed with cancer or severe hematological disorder compared to healthy controls and this reduction is disease and age dependent: patients with central nervous system cancer (CNS tumors) and hematological disorders, as well as boys <7 years are the most affected. WHAT IS KNOWN ALREADY: Fertility preservation in pediatric male patients is considered based on the gonadotoxicity of selected treatments. Although treatment effects on germ cells have been extensively investigated, limited data are available on the effect of the disease on the prepubertal male gonad. Of the few studies investigating the effects of cancer or hematologic disorders on testicular function and germ cell quantity in prepuberty, the results are inconsistent. However, recent studies suggested impairments before the initiation of known gonadotoxic therapy. Understanding which diseases and at what age affect the germ cell pool in pediatric patients before treatment is critical to optimize strategies and counseling for fertility preservation. STUDY DESIGN, SIZE, DURATION: This multicenter retrospective cohort study included 101 boys aged <14 years with extra-cerebral cancer (solid tumors), CNS tumors, leukemia/lymphoma (blood cancer), or non-malignant hematological disorders, who were admitted for a fertility preservation programme between 2002 and 2018. PARTICIPANTS/MATERIALS, SETTING, METHODS: In addition to clinical data, we analyzed measurements of testicular volume and performed histological staining on testicular biopsies obtained before treatment, at cryopreservation, to evaluate number of spermatogonia per tubular cross-section, tubular fertility index, and the most advanced germ cell type prior to chemo-/radiotherapy. The controls were data simulations with summary statistics from original studies reporting healthy prepubertal boys' testes characteristics. MAIN RESULTS AND THE ROLE OF CHANCE: Prepubertal patients with childhood cancer or hematological disorders were more likely to have significantly reduced spermatogonial quantity compared to healthy controls (48.5% versus 31.0% prevalence, respectively). The prevalence of patients with reduced spermatogonial quantity was highest in the CNS tumor (56.7%) and the hematological disorder (55.6%) groups, including patients with hydroxyurea pre-treated sickle cell disease (58.3%) and patients not exposed to hydroxyurea (50%). Disease also adversely impacted spermatogonial distribution and differentiation. Irrespective of disease, we observed the highest spermatogonial quantity reduction in patients <7 years of age. LIMITATIONS, REASONS FOR CAUTION: For ethical reasons, we could not collect spermatogonial quantity data in healthy prepubertal boys as controls and thus deployed statistical simulation on data from literature. Also, our results should be interpreted considering low patient numbers per (sub)group. WIDER IMPLICATIONS OF THE FINDINGS: Cancers, especially CNS tumors, and severe hematological disorders can affect spermatogonial quantity in prepubertal boys before treatment. Consequently, these patients may have a higher risk of depleted spermatogonia following therapies, resulting in persistent infertility. Therefore, patient counseling prior to disease treatment and timing of fertility preservation should not only be based on treatment regimes, but also on diagnoses and age. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by Marie Curie Initial Training Network (ITN) (EU-FP7-PEOPLE-2013-ITN) funded by European Commision grant no. 603568; ZonMW Translational Adult stem cell research (TAS) grant no. 116003002. No competing interests. TRIAL REGISTRATION NUMBER: N/A.


Asunto(s)
Preservación de la Fertilidad , Enfermedades Hematológicas , Neoplasias , Adulto , Niño , Humanos , Masculino , Espermatogonias , Preservación de la Fertilidad/métodos , Estudios Retrospectivos , Hidroxiurea , Testículo , Criopreservación
2.
Pediatr Blood Cancer ; 70(8): e30370, 2023 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-37150973

RESUMEN

BACKGROUND: Experimental fertility preservation programs have been started to safeguard the future fertility of prepubertal and pubertal males requiring high-risk gonadotoxic treatment protocols. However, long-term follow-up studies evaluating the effects on their gonadal development and function related to the testicular biopsy procedure are rather limited. DESIGN: This two-center follow-up study (between 2002 and 2020) evaluated the gonadal development and function of a cohort of 59 prepubertal and pubertal males who have been offered immature testicular tissue banking (TTB) prior to conventional high-risk chemo- and/or radiotherapy (HR-C/R) or conditioning therapy before hematopoietic stem cell transplantation (CT-HSCT). The aim is to investigate the long-term impact of the testicular biopsy procedure and the high-risk gonadotoxic treatment. Testicular growth and the reproductive hormones luteinizing hormone (LH), follicle-stimulating hormone (FSH), testosterone (T), and inhibin B (INHB) were analyzed after treatment completion, and compared between males accepting TTB and those refusing TTB (control) as well as between HR-C/R and CT-HSCT treatment protocols. RESULTS: Of the 59 prepubertal and pubertal males included, 25 were treated by HR-C/R and 34 required CT-HSCT. TTB was accepted for 39 males and refused for 20 males. Most patients were prepubertal at diagnosis (85%), at TTB (79%), and at treatment completion (76%), and pubertal or postpubertal at their last follow-up visit (66%). After 5.0 (1.0-13.0) years post treatment, most patients show normal testicular volumes (83%) and normal LH (89%), FSH (87%), T (87%), and INHB (79%) serum levels. The testicular biopsy procedure did not have an effect on testicular growth, LH, FSH, T, and INHB. Significantly more small postpubertal testicular volumes (p = .0278) and low INHB serum levels (p = .0130) were recorded after CT-HSCT, especially after myeloablative conditioning. CONCLUSION: The clinical follow-up data demonstrate no effect related to the biopsy procedure, but a substantial risk for impaired gonadal development after high-risk gonadotoxic treatment, in particular myeloablative CT-HSCT. Longer follow-up studies with a larger study population are needed to confirm these preliminary findings.


Asunto(s)
Hormona Luteinizante , Testículo , Masculino , Humanos , Estudios de Seguimiento , Hormona Folículo Estimulante , Testosterona
3.
Reprod Biomed Online ; 44(5): 889-895, 2022 05.
Artículo en Inglés | MEDLINE | ID: mdl-35282994

RESUMEN

RESEARCH QUESTION: Which early-diagnosed Klinefelter syndrome patients have been offered cryopreservation of testicular tissue as part of fertility preservation before spermatogonial stem cell (SSC) loss? Do these Klinefelter syndrome patients present with behavioural, cognitive and/or psychological problems? Does a testicular biopsy procedure have long-term effects on the gonadal development of Klinefelter syndrome patients? DESIGN: Early-diagnosed Klinefelter syndrome patients followed between 2009 and 2020 and offered testicular tissue banking in an experimental context at the Universitair Ziekenhuis Brussel were included. The prevalence of behavioural, cognitive and/or psychological problems was determined. Changes in testicular volume and in gonadal function (LH, FSH, testosterone and inhibin B [INHB]) were studied. RESULTS: Of the 48 Klinefelter syndrome patients included, 22 had testicular tissue removed (biopsy group) and 26 had no surgical intervention (control group). The need for specialized education was significantly higher in prenatally (P = 0.0159) and prepubertally (P = 0.0002) diagnosed Klinefelter syndrome patients. Psychological problems were significantly more prevalent in Klinefelter syndrome patients who did not opt for fertility preservation (P = 0.0447). In the first 4.2 (1.9-9.1) years after testicular biopsy, no difference in testicular volume was observed between the biopsied and the contralateral non-biopsied testes (P > 0.9999). After pubertal onset, no differences in LH, FSH, testosterone and INHB were found between the biopsy and the control groups (P = 0.1324 for LH, P > 0.9999 for FSH, P = 0.5433 for testosterone, P > 0.9999 for INHB). CONCLUSION: Early-diagnosed Klinefelter syndrome patients presented with behavioural, cognitive and/or psychological problems. Only psychological problems seemed to influence the decision towards fertility preservation. Follow-up data confirm that harvesting testicular tissue does not have a long-term impact on the gonadal development of Klinefelter syndrome patients.


Asunto(s)
Preservación de la Fertilidad , Síndrome de Klinefelter , Biopsia , Femenino , Preservación de la Fertilidad/métodos , Hormona Folículo Estimulante , Estudios de Seguimiento , Humanos , Síndrome de Klinefelter/complicaciones , Síndrome de Klinefelter/diagnóstico , Síndrome de Klinefelter/patología , Masculino , Testículo/patología , Testosterona
4.
Reprod Biomed Online ; 44(5): 896-906, 2022 05.
Artículo en Inglés | MEDLINE | ID: mdl-35282995

RESUMEN

RESEARCH QUESTION: Is intratesticular xenotransplantation a potential ex-vivo model for studying testicular fibrosis related to Klinefelter syndrome? STUDY DESIGN: First, a feasibility study of an ex-vivo model to study testicular fibrosis in patients with Klinefelter syndrome was performed. Testis tissue from boys with pre-pubertal Klinefelter syndrome (n = 3) and controls (n = 2) (<18 years) was grafted to the mouse testis (n = 12) and recovered after 2, 4, 6 and 8 weeks. Part two of this study consisted of a validation of this model, evaluating the effects of the mast cell blocker ketotifen on the histology of the grafts of Klinefelter syndrome (n = 5) and controls (n = 3), transplanted to mice (n = 10), after 4 weeks of ketotifen or saline treatment. Immunohistochemistry determined the histology of the grafts and the presence of mast cells and spermatogonia. RESULTS: The feasibility study showed that 4 weeks after transplantation, all Klinefelter syndrome grafts could be recovered. Later, degeneration was observed. Most recovered grafts showed an intact histology, with 67 ± 12% intact tubules for the Klinefelter syndrome grafts and 65 ± 15% of intact tubules for the control grafts. In the few remaining Klinefelter syndrome grafts, treatment with ketotifen improved testicular histology compared with non-treated grafts. Graft survival was patient dependent. No germ cell loss was observed after transplantation. CONCLUSION: Xenografting could become a model for the longitudinal study of the fibrotic process related to Klinefelter syndrome; however, the current model has a limited survival period and patient-specific differences in histology.


Asunto(s)
Síndrome de Klinefelter , Testículo , Animales , Femenino , Fibrosis , Humanos , Cetotifen , Síndrome de Klinefelter/patología , Estudios Longitudinales , Masculino , Ratones , Espermatogénesis , Espermatogonias , Testículo/patología , Trasplante Heterólogo
5.
Hum Reprod ; 36(1): 5-15, 2021 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-33257947

RESUMEN

STUDY QUESTION: Does gender-affirming treatment prevent full spermatogenesis in transgender women (TW)? SUMMARY ANSWER: Adequate hormonal therapy (HT) leads to complete suppression of spermatogenesis in most TW, if serum testosterone levels within female reference ranges are obtained. WHAT IS KNOWN ALREADY: Gender-affirming treatment in transgender individuals may involve gender-affirming HT. The effects on spermatogenesis in TW remain unclear. In order to add information from a referral centre for transgender care, we wish to compare results of earlier studies with our population of TW who received a standard hormone treatment. STUDY DESIGN, SIZE, DURATION: This was a prospective cohort study part of the European Network for the Investigation of Gender Incongruence (ENIGI), conducted between 15 February 2010 and 30 September 2015. There were 162 TW were included in the ENIGI study at the Ghent University Hospital in Belgium. Participants are included in ENIGI when they first start HT, and follow-up visits occur over the next 3 years. PARTICIPANTS/MATERIALS, SETTING METHODS: The study included 97 TW who initiated HT with cyproterone acetate (CPA) plus oestrogens and proceeded with gonadectomy at the Ghent University Hospital. Testicular tissue retrieved during gonadectomy was processed and stained for four different germ cell markers by the Biology of the Testis lab at the Vrije Universiteit Brussel. Subsequent immunohistochemical staining was performed for melanoma-associated antigen A4 (MAGE-A4, marker for spermatogonia and early spermatocytes), boule homologue, RNA-binding protein (BOLL, marker for secondary spermatocytes and round spermatids), cAMP-responsive element modulator (CREM, marker for round spermatids) and acrosin (marker for acrosome visualization). Serum levels of sex steroids were measured prior to surgery. MAIN RESULTS AND THE ROLE OF CHANCE: Suppressed testosterone levels (<50 ng/dl) were found in 92% of the participants prior to surgery. The mean time between initiation of HT and surgery was 685 days. In 88% (85/97) of the sections, MAGE-A4 staining was positive. Further staining could not reveal complete spermatogenesis in any participant. LIMITATIONS, REASONS FOR CAUTION: Testicular function of the participants prior to initiation of HT was not assessed, although all participants presented with cisgender male serum testosterone values before initiation of HT. The current study only reports on people using CPA at a fixed dose and may therefore not be applicable to all TW. WIDER IMPLICATIONS OF THE FINDINGS: HT leads to complete suppression of spermatogenesis in most TW, if serum testosterone levels within female reference ranges are obtained. Serum testosterone levels are associated with the sperm maturation rate. It is important to discuss sperm preservation before the start of hormone therapy. If serum testosterone levels remain higher, spermatogenesis may still occur. STUDY FUNDING/COMPETING INTEREST(S): D.V.S. is a post-doctoral fellow of the Fonds Wetenschappelijk Onderzoek (FWO; 12M2819N). Processing of the testis specimens was funded by the Biology of The Testes (BITE) research group (Department of Reproduction, Genetics and Regenerative medicine at Vrije Universiteit Brussel (VUB)). There are no competing interests. TRIAL REGISTRATION NUMBER: N/A.


Asunto(s)
Personas Transgénero , Bélgica , Femenino , Humanos , Masculino , Estudios Prospectivos , Espermatogénesis , Espermatogonias , Testículo
6.
J Inherit Metab Dis ; 44(6): 1393-1408, 2021 11.
Artículo en Inglés | MEDLINE | ID: mdl-34494673

RESUMEN

Cystinosis is an inherited metabolic disorder caused by autosomal recessive mutations in the CTNS gene leading to lysosomal cystine accumulation. The disease primarily affects the kidneys followed by extra-renal organ involvement later in life. Azoospermia is one of the unclarified complications which are not improved by cysteamine, which is the only available disease-modifying treatment. We aimed at unraveling the origin of azoospermia in cysteamine-treated cystinosis by confirming or excluding an obstructive factor, and investigating the effect of cysteamine on fertility in the Ctns-/- mouse model compared with wild type. Azoospermia was present in the vast majority of infantile type cystinosis patients. While spermatogenesis was intact, an enlarged caput epididymis and reduced levels of seminal markers for obstruction neutral α-glucosidase (NAG) and extracellular matrix protein 1 (ECM1) pointed towards an epididymal obstruction. Histopathological examination in human and mouse testis revealed a disturbed blood-testis barrier characterized by an altered zonula occludens-1 (ZO-1) protein expression. Animal studies ruled out a negative effect of cysteamine on fertility, but showed that cystine accumulation in the testis is irresponsive to regular cysteamine treatment. We conclude that the azoospermia in infantile cystinosis is due to an obstruction related to epididymal dysfunction, irrespective of the severity of an evolving primary hypogonadism. Regular cysteamine treatment does not affect fertility but has subtherapeutic effects on cystine accumulation in testis.


Asunto(s)
Azoospermia/patología , Barrera Hematotesticular/metabolismo , Cisteamina/uso terapéutico , Cistinosis/tratamiento farmacológico , Testículo/patología , Adulto , Animales , Azoospermia/complicaciones , Azoospermia/genética , Depletores de Cistina/uso terapéutico , Cistinosis/complicaciones , Cistinosis/patología , Modelos Animales de Enfermedad , Proteínas de la Matriz Extracelular/metabolismo , Humanos , Infertilidad Masculina/etiología , Infertilidad Masculina/genética , Infertilidad Masculina/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Proteínas de Neoplasias/metabolismo , Estudios Retrospectivos , Adulto Joven , Proteína de la Zonula Occludens-1/metabolismo
7.
Hum Reprod ; 35(8): 1753-1764, 2020 08 01.
Artículo en Inglés | MEDLINE | ID: mdl-32649748

RESUMEN

STUDY QUESTION: Is the distribution of immune cells and the testicular vasculature altered in testicular biopsies from patients with Klinefelter syndrome (KS)? SUMMARY ANSWER: Increased numbers of macrophages and mast cells, an increased expression of decorin and an increased blood vessel density were found in KS samples compared to controls. WHAT IS KNOWN ALREADY: Most KS patients are infertile due to an early germ cell loss. From puberty onwards, testicular fibrosis can be detected. How this fibrotic process is initiated remains unknown. STUDY DESIGN, SIZE, DURATION: In this study, the number of macrophages, mast cells and their secretory products were evaluated in KS, Sertoli cell only (SCO) and control patient samples. The association between immune cell numbers and level of fibrosis in KS tissue was examined. In addition, the vascularization within these testicular tissue biopsies was studied. For immunohistochemical evaluation, KS patients at different stages of testicular development were included: prepubertal (aged 4-7 years; n = 4), peripubertal (aged 11-17 years; n = 21) and adult (aged >18 years; n = 37) patients. In addition, testicular tissue biopsies of adult SCO (n = 33) and control samples for the three KS age groups (prepubertal n = 9; peripubertal n = 5; adult n = 25) were analysed. Gene expression analysis was performed on adult testicular tissue from KS (n = 5), SCO (n = 5) and control (n = 5) patients. PARTICIPANTS/MATERIALS, SETTING, METHODS: Adult (>18 years) KS, SCO and control testicular tissue biopsies were obtained during a testicular sperm extraction procedure. KS peripubertal (11-18 years), prepubertal (<11 years) and age-matched control biopsies were obtained from the biobank of the university hospital. Immunohistochemistry was used to determine the tubular structure (H/PAS), the number of spermatogonia (MAGE-A4), macrophages (CD68) and mast cells (tryptase) and the blood vessel density (Von Willebrand factor). In addition, quantitative real-time polymerase chain reaction was used to determine the expression of secretory products of macrophages and mast cells (tryptase, tumour necrosis factor alpha and decorin). MAIN RESULTS AND THE ROLE OF CHANCE: A significant increase in the number of macrophages (P < 0.0001) and mast cells (P = 0.0008) was found in the peritubular compartment of testes of adult KS patients compared to control samples. However, no association between the number of immune cells and the degree of fibrosis was observed. In adult SCO samples, a significant increase was seen for peritubular macrophage (P < 0.0001) and mast cell (P < 0.0001) numbers compared to control samples. In the interstitial compartment, a significant increase in mast cell number was found in adult SCO samples compared to KS (P < 0.0001) and control (P < 0.0001) tissue. A significant difference (P = 0.0431) in decorin expression could be detected in adult KS compared to control patients. Decorin expression was mostly seen in the walls of the seminiferous tubules. When comparing the vascularization between KS patients and age-matched controls, a significant increase (P = 0.0081) in blood vessel density could be observed only in prepubertal KS testicular tissue. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: As controls for this study, testicular tissue biopsies of men who underwent a vasectomy reversal or orchiectomy were used, but these men may not represent fertile controls. In addition, a high variability in immune cell numbers, secretory products expression and number of blood vessels could be observed amongst all patient samples. WIDER IMPLICATIONS OF THE FINDINGS: Increased numbers of macrophages and mast cells have previously been described in non-KS infertile men. Our results show that these increased numbers can also be detected in KS testicular tissue. However, no association between the number of macrophages or mast cells and the degree of fibrosis in KS samples could be detected. Decorin has previously been described in relation to fibrosis, but it has not yet been associated with testicular fibrosis in KS. Our results suggest a role for this proteoglycan in the fibrotic process since an increased expression was observed in adult KS tissue compared to controls. Impaired vascularization in KS men was suggested to be responsible for the KS-related disturbed hormone levels. Our results show a significant difference in blood vessel density, especially for the smallest blood vessels, between prepubertal KS samples and age-matched controls. This is the first study to report differences between KS and control testicular tissue at prepubertal age. STUDY FUNDING/COMPETING INTEREST(S): The project was funded by grants from the Vrije Universiteit Brussel (E.G.) and the scientific Fund Willy Gepts from the UZ Brussel (D.V.S.). D.V.S. is a post-doctoral fellow of the Fonds voor Wetenschappelijk Onderzoek (FWO; 12M2819N). No conflict of interest is declared for this research project.


Asunto(s)
Síndrome de Klinefelter , Testículo , Adolescente , Adulto , Niño , Preescolar , Humanos , Síndrome de Klinefelter/genética , Masculino , Túbulos Seminíferos , Células de Sertoli , Espermatogonias
8.
Reprod Biomed Online ; 40(2): 261-269, 2020 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-32001160

RESUMEN

RESEARCH QUESTION: Which cryopreservation method better protects reproductive potential: the cryopreservation of a testicular cell suspension (TCS) or the cryopreservation of testicular tissue (TET)? DESIGN: Two cryopreservation strategies for spermatogonial stem cells (SSCs) were compared in a mouse model: cryopreservation as TET or as TCS. Evaluated outcomes were number of viable cells after thawing, number and length of donor-derived colonies after spermatogonial stem cell transplantation (SSCT), number of litters, litter size and number of donor-derived pups after mating. RESULTS: Compared with cryopreserving TCS, cryopreservation of TET resulted in significantly higher numbers of viable cells after thawing (TET: 13.4 â€¯×  104 ± 7.2 â€¯×  104 versus TCS: 8.2 â€¯×  104 ± 2.7 â€¯×  104; P = 0.0002), more (TET: 47.6 ± 19.2 versus TCS: 18.5 ± 13.0; P = 0.0039) and longer (TET: 5.2 ± 1.0 mm versus TCS: 2.7 ± 1.5 mm; P = 0.0016) donor-derived colonies, and more donor-derived pups per litter (TET: 2.2 ± 0.2 versus TCS: 0.5 ± 0.1; P = 0.0008). CONCLUSIONS: Cryopreservation of TET is the preferred method to cryopreserve SSCs prior to SSCT in a mouse model.


Asunto(s)
Células Madre Germinales Adultas , Preservación de la Fertilidad/métodos , Fertilidad/fisiología , Testículo/trasplante , Animales , Criopreservación , Masculino , Ratones
9.
Reprod Biomed Online ; 39(1): 119-133, 2019 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-31005581

RESUMEN

RESEARCH QUESTION: Does recombinant human vascular endothelial growth factor (VEGF-165) improve the efficiency of human immature testis tissue (ITT) xenotransplantation? DESIGN: ITT fragments from three prepubertal boys were cultured for 5 days with VEGF-165 or without (control) before xenotransplantation into the testes of immunodeficient mice. Xenotransplants were recovered at 4 and 9 months post-transplantation and vascularization, seminiferous tubule integrity, number of spermatogonia and germ cell differentiation were evaluated by histology and immunohistochemistry. RESULTS: Transplants from donor 1 and donor 2 treated with VEGF demonstrated higher vascular surface (P = 0.004) and vessel density (P = 0.011) overall and contained more intact seminiferous tubules (P = 0.039) with time, compared with controls. The number of spermatogonia was increased over time (P < 0.001) irrespective of treatment and donor, whereas, for the VEGF-treated transplants, the increase was even higher over time (P = 0.020). At 9 months, spermatocytes were present in the xenotransplants, irrespective of treatment. No transplants could be recovered from donor 3, who had already received treatment with cyclosporine for aplastic anaemia before biopsy. CONCLUSIONS: In-vitro pre-treatment of human prepubertal testis tissue with VEGF improved transplant vascularization in two out of three cases, resulting in improved seminiferous tubule integrity and spermatogonial survival during xenotransplantation. Although further studies are warranted, we suggest VEGF to be considered as a factor for improving the efficiency of immature testis tissue transplantation in the future.


Asunto(s)
Testículo/efectos de los fármacos , Testículo/trasplante , Trasplante Heterólogo , Factor A de Crecimiento Endotelial Vascular/farmacología , Factores de Edad , Animales , Biopsia , Diferenciación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Niño , Preescolar , Criopreservación , Preservación de la Fertilidad/métodos , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Desnudos , Ratones Transgénicos , Pubertad/fisiología , Proteínas Recombinantes/farmacología , Espermatogénesis/efectos de los fármacos , Espermatogonias/citología , Espermatogonias/efectos de los fármacos , Espermatogonias/fisiología , Testículo/citología , Testículo/patología
10.
Int J Mol Sci ; 20(20)2019 Oct 21.
Artículo en Inglés | MEDLINE | ID: mdl-31640294

RESUMEN

While the incidence of cancer in children and adolescents has significantly increased over the last decades, improvements made in the field of cancer therapy have led to an increased life expectancy for childhood cancer survivors. However, the gonadotoxic effect of the treatments may lead to infertility. Although semen cryopreservation represents the most efficient and safe fertility preservation method for males producing sperm, it is not feasible for prepubertal boys. The development of an effective strategy based on the pharmacological protection of the germ cells and testicular function during gonadotoxic exposure is a non-invasive preventive approach that prepubertal boys could benefit from. However, the progress in this field is slow. Currently, cryopreservation of immature testicular tissue (ITT) containing spermatogonial stem cells is offered to prepubertal boys as an experimental fertility preservation strategy by a number of medical centers. Several in vitro and in vivo fertility restoration approaches based on the use of ITT have been developed so far with autotransplantation of ITT appearing more promising. In this review, we discuss the pharmacological approaches for fertility protection in prepubertal and adolescent boys and the fertility restoration approaches developed on the utilization of ITT.


Asunto(s)
Células Madre Germinales Adultas/citología , Neoplasias/complicaciones , Testículo/trasplante , Adolescente , Células Madre Germinales Adultas/química , Niño , Preescolar , Criopreservación , Preservación de la Fertilidad , Humanos , Lactante , Masculino , MicroARNs/genética , Neoplasias/terapia , Conservación de Tejido
11.
Hum Reprod ; 33(9): 1581-1585, 2018 09 01.
Artículo en Inglés | MEDLINE | ID: mdl-30020439

RESUMEN

It is widely acknowledged that the responsible introduction of new assisted reproductive technologies (ARTs) requires preclinical safety research, including the use of animal models and human embryos. However, the moral sensitivity of human embryo research has led to regulations and guidance stating that human embryos may only be used for research that cannot also be conducted with animals. We call this the 'use animals first' (UAF) rule. In the field of ART research, this translates into the notion of an ideal chain of consecutive preclinical research steps, where research using human embryos may only be considered as a further step after promising results have been obtained in animals first. This may lead to research ethics committees requiring animal studies that are in fact a waste of time and money, while exposing animals to an infringement of their wellbeing for no good purpose. In this paper, we explore the possible moral arguments behind the UAF-rule and test their validity. We conclude that there are no convincing grounds for upholding this rule and recommend replacing it.


Asunto(s)
Técnicas Reproductivas Asistidas/ética , Creación de Embriones para Investigación/ética , Animales , Comités de Ética en Investigación , Femenino , Humanos , Modelos Animales , Donación de Oocito/efectos adversos , Donación de Oocito/ética , Creación de Embriones para Investigación/legislación & jurisprudencia
12.
Reprod Biomed Online ; 37(1): 6-17, 2018 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-29776850

RESUMEN

RESEARCH QUESTION: From a clinical perspective, which parameters grant optimal cryopreservation of mouse testicular cell suspensions? DESIGN: We studied the effect of different cryopreservation rates, the addition of sugars, different vessels and the addition of an apoptotic inhibitor on the efficiency of testicular cell suspension cryopreservation. After thawing and warming, testicular cell suspensions were transplanted to recipient mice for further functional assay. After selecting the optimal cryopreservation procedure, a second experiment compared the transplantation efficiency between the selected freezing protocol and fresh testicular cell suspensions. RESULTS: Multiple- and single-step freezing did not differ significantly in terms of recovered viable cells (RVC) (33 ± 28% and 38 ± 25%). The addition of sucrose did not result in a higher RVC (33 ± 20%). Cells frozen in vials recovered better than those frozen in straws (52 ± 20% versus 33 ± 20%; P = 0.0049). The inclusion of an apoptosis inhibitor (z-VAD[Oe]-FMK) significantly increased the RVC after thawing (61 ± 18% versus 50 ± 17%; P = 0.0480). When comparing the optimal cryopreservation procedure with fresh testicular cell suspensions, a lower RVC (63 ± 11% versus 92 ± 4%; P < 0.0001) and number of donor-derived spermatogonial stem cell colonies per testis (34.04 ± 2.34 versus 16.78 ± 7.76; P = 0.0051) were observed. CONCLUSION: Upon freeze-thawing or vitrification-warming, and assessment of donor-derived spermatogenesis after transplantation, Dulbecco's modified Eagle's medium supplemented with 1.5M dimethyl-sulphoxide, 10% fetal calf serum and 60 µM of Z-VAD-(OMe)-FMK in vials at a freezing rate of -1°C/min was optimal.


Asunto(s)
Criopreservación/métodos , Crioprotectores/farmacología , Espermatogénesis/efectos de los fármacos , Testículo/citología , Animales , Masculino , Ratones , Testículo/efectos de los fármacos , Vitrificación
13.
Med Health Care Philos ; 21(4): 537-545, 2018 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-29417302

RESUMEN

In the field of medically assisted reproduction (MAR), there is a growing emphasis on the importance of introducing new assisted reproductive technologies (ARTs) only after thorough preclinical safety research, including the use of animal models. At the same time, there is international support for the three R's (replace, reduce, refine), and the European Union even aims at the full replacement of animals for research. The apparent tension between these two trends underlines the urgency of an explicit justification of the use of animals for the development and preclinical testing of new ARTs. Considering that the use of animals remains necessary for specific forms of ART research and taking account of different views on the moral importance of helping people to have a genetically related child, we argue that, in principle, the importance of safety research as part of responsible innovation outweighs the limited infringement of animal wellbeing involved in ART research.


Asunto(s)
Experimentación Animal/ética , Bienestar del Animal/ética , Investigación Biomédica/ética , Investigación Biomédica/métodos , Técnicas Reproductivas Asistidas , Animales , Humanos , Principios Morales , Políticas
14.
Hum Mol Genet ; 24(19): 5581-8, 2015 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-26199321

RESUMEN

Infertility is a global healthcare problem, and despite long years of assisted reproductive activities, a significant number of cases remain idiopathic. Our currently restricted understanding of basic mechanisms driving human gametogenesis severely limits the improvement of clinical care for infertile patients. Using exome sequencing, we identified a nonsense mutation leading to a premature stop in the TEX15 locus (c.2130T>G, p.Y710*) in a consanguineous Turkish family comprising eight siblings in which three brothers were identified as infertile. TEX15 displays testis-specific expression, maps to chromosome 8, contains four exons and encodes a 2789-amino acid protein with uncertain function. The mutation, which should lead to early translational termination at the first exon of TEX15, co-segregated with the infertility phenotype, and our data strongly suggest that it is the cause of spermatogenic defects in the family. All three affected brothers presented a phenotype reminiscent of the one observed in KO mice. Indeed, previously reported results demonstrated that disruption of the orthologous gene in mice caused a drastic reduction in testis size and meiotic arrest in the first wave of spermatogenesis in males while female KO mice were fertile. The data from our study of one Turkish family suggested that the identified mutation correlates with a decrease in sperm count over time. A diagnostic test identifying the mutation in man could provide an indication of spermatogenic failure and prompt patients to undertake sperm cryopreservation at an early age.


Asunto(s)
Proteínas de Ciclo Celular/genética , Codón sin Sentido , Infertilidad Masculina/genética , Análisis de Secuencia de ADN/métodos , Espermatogénesis , Población Blanca/genética , Consanguinidad , Exoma , Predisposición Genética a la Enfermedad , Humanos , Infertilidad Masculina/patología , Masculino , Meiosis , Oligospermia , Tamaño de los Órganos , Linaje , Testículo/anatomía & histología , Factores de Tiempo , Turquía
15.
Reprod Biomed Online ; 35(2): 180-188, 2017 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-28583375

RESUMEN

During short-term storage, before cryopreservation, testicular tissue quality can be affected by storage medium, duration, temperature and tissue size. We previously established the best storage medium and time for short-term maintenance of tissue. In this study, three different storage temperatures (4°C, room temperature, 37°C) and four tissue sizes (~6 mm3; ~ 15 mm3; ~ 50 mm3 or ~80 mm3) were assessed over the course of a fixed period of 3 days. Storing human testicular tissue at 37°C caused a significant increase in the number of apoptotic cells per tubule (P = 0.002), compared with fresh control, but this was not the case at 4°C or room temperature. Temperature did not affect viability, tissue morphology or number of spermatogonia in samples. The morphology of the testicular tissue was optimally preserved when stored as large fragments (~50 mm3: P = 0.018; ~ 80 mm3: P = 0.018). Tissue size did not significantly affect viability, number of spermatogonia or apoptotic cells. Adult human testicular tissue can be preserved at 4°C or room temperature without altering tissue morphology, Sertoli cell morphology, number of spermatogonia or number of apoptotic cells. The tissue does not need to be extensively dissected as tissue morphology is better maintained in larger fragments.


Asunto(s)
Criopreservación , Testículo , Humanos , Masculino , Células de Sertoli/citología , Espermatogonias/citología , Temperatura
16.
Reprod Biomed Online ; 34(3): 291-297, 2017 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-28040413

RESUMEN

When fertility is impaired by anticancer treatment, spermatogonial stem cell transplantation (SSCT) could be used as a fertility restoration technique later on in life. Previously, we have demonstrated that a testicular cell suspension could be injected into a human cadaver testis, however, leakage to the interstitium was observed. In this study, injection of mouse testicular cells at an injection height of 50 cm (hydrostatic pressure) or via an automated injection pump (1400 µl, 2600 µl and 3000 µl) was evaluated. Significant difference in the filled radioactive volume was reached between the group in which 1400 µl was injected with an infusion pump and the groups in which 2600 µl (P = 0.019) or 3000 µl (P = 0.010) was injected. In all experimental groups green fluorescent protein positive (GFP+) cells were observed in the seminiferous tubules. In conclusion, a lower injection height did not resolve the leakage of the injected cells to the interstitium. Using the infusion pump resulted in more efficient filling of the seminiferous tubules with lower interexperimental variability. Although leakage to the interstitium was still observed, with further optimisation, the use of an infusion pump for clinical application is advantageous.


Asunto(s)
Células Madre Germinales Adultas/trasplante , Trasplante de Células Madre/métodos , Animales , Preservación de la Fertilidad/métodos , Masculino , Ratones , Túbulos Seminíferos/citología
17.
J Assist Reprod Genet ; 34(5): 683-694, 2017 May.
Artículo en Inglés | MEDLINE | ID: mdl-28401488

RESUMEN

PURPOSE: The purpose of this study was to identify mutations that cause non-syndromic male infertility using whole exome sequencing of family cases. METHODS: We recruited a consanguineous Turkish family comprising nine siblings with male triplets; two of the triplets were infertile as well as one younger infertile brother. Whole exome sequencing (WES) performed on two azoospermic brothers identified a mutation in the melanoma antigen family B4 (MAGEB4) gene which was confirmed via Sanger sequencing and then screened for on control groups and unrelated infertile subjects. The effect of the mutation on messenger RNA (mRNA) and protein levels was tested after in vitro cell transfection. Structural features of MAGEB4 were predicted throughout the conserved MAGE domain. RESULTS: The novel single-base substitution (c.1041A>T) in the X-linked MAGEB4 gene was identified as a no-stop mutation. The mutation is predicted to add 24 amino acids to the C-terminus of MAGEB4. Our functional studies were unable to detect any effect either on mRNA stability, intracellular localization of the protein, or the ability to homodimerize/heterodimerize with other MAGE proteins. We thus hypothesize that these additional amino acids may affect the proper protein interactions with MAGEB4 partners. CONCLUSION: The whole exome analysis of a consanguineous Turkish family revealed MAGEB4 as a possible new X-linked cause of inherited male infertility. This study provides the first clue to the physiological function of a MAGE protein.


Asunto(s)
Antígenos de Neoplasias/genética , Azoospermia/genética , Genes Ligados a X/genética , Infertilidad Masculina/genética , Proteínas de Neoplasias/genética , Oligospermia/genética , Adulto , Azoospermia/patología , Preescolar , Consanguinidad , Frecuencia de los Genes , Homocigoto , Humanos , Infertilidad Masculina/patología , Masculino , Mutación , Oligospermia/patología , Linaje , Polimorfismo de Nucleótido Simple , ARN Mensajero/genética , Secuenciación del Exoma
18.
Hum Reprod ; 30(11): 2463-75, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-26358785

RESUMEN

STUDY QUESTION: What clinical practices, patient management strategies and experimental methods are currently being used to preserve and restore the fertility of prepubertal boys and adolescent males? SUMMARY ANSWER: Based on a review of the clinical literature and research evidence for sperm freezing and testicular tissue cryopreservation, and after consideration of the relevant ethical and legal challenges, an algorithm for the cryopreservation of sperm and testicular tissue is proposed for prepubertal boys and adolescent males at high risk of fertility loss. WHAT IS KNOWN ALREADY: A known late effect of the chemotherapy agents and radiation exposure regimes used to treat childhood cancers and other non-malignant conditions in males is the damage and/or loss of the proliferating spermatogonial stem cells in the testis. Cryopreservation of spermatozoa is the first line treatment for fertility preservation in adolescent males. Where sperm retrieval is impossible, such as in prepubertal boys, or it is unfeasible in adolescents prior to the onset of ablative therapies, alternative experimental treatments such as testicular tissue cryopreservation and the harvesting and banking of isolated spermatogonial stem cells can now be proposed as viable means of preserving fertility. STUDY DESIGN, SIZE, DURATION: Advances in clinical treatments, patient management strategies and the research methods used to preserve sperm and testicular tissue for prepubertal boys and adolescents were reviewed. A snapshot of the up-take of testis cryopreservation as a means to preserve the fertility of young males prior to December 2012 was provided using a questionnaire. PARTICIPANTS/MATERIALS, SETTING, METHODS: A comprehensive literature review was conducted. In addition, survey results of testis freezing practices in young patients were collated from 24 European centres and Israeli University Hospitals. MAIN RESULTS AND THE ROLE OF CHANCE: There is increasing evidence of the use of testicular tissue cryopreservation as a means to preserve the fertility of pre- and peri-pubertal boys of up to 16 year-old. The survey results indicate that of the 14 respondents, half of the centres were actively offering testis tissue cryobanking as a means of safeguarding the future fertility of boys and adolescents as more than 260 young patients (age range less than 1 year old to 16 years of age), had already undergone testicular tissue retrieval and storage for fertility preservation. The remaining centres were considering the implementation of a tissue-based fertility preservation programme for boys undergoing oncological treatments. LIMITATIONS, REASONS FOR CAUTION: The data collected were limited by the scope of the questionnaire, the geographical range of the survey area, and the small number of respondents. WIDER IMPLICATIONS OF THE FINDINGS: The clinical and research questions identified and the ethical and legal issues raised are highly relevant to the multi-disciplinary teams developing treatment strategies to preserve the fertility of prepubertal and adolescent boys who have a high risk of fertility loss due to ablative interventions, trauma or genetic pre-disposition.


Asunto(s)
Criopreservación/métodos , Preservación de la Fertilidad/métodos , Testículo , Adolescente , Niño , Europa (Continente) , Humanos , Masculino
19.
Cancers (Basel) ; 16(6)2024 Mar 17.
Artículo en Inglés | MEDLINE | ID: mdl-38539511

RESUMEN

In recent years, immune checkpoint inhibitors (ICIs) have become a viable option for many cancer patients, including specific subgroups of pediatric patients. Despite their efficiency in treating different types of cancer, ICIs are responsible for a number of immune-related adverse events, including inflammatory toxicities, that can affect several organs. However, our knowledge of the impact of ICIs on the testis and male fertility is limited. It is possible that ICI treatment affects testicular function and spermatogenesis either directly or indirectly (or both). Treatment with ICIs may cause increased inflammation and immune cell infiltration within the seminiferous tubules of the testis, disturbing spermatogenesis or testosterone deficiency (primary hypogonadism). Additionally, the interference of ICIs with the hypothalamic-pituitary-gonadal axis may alter testosterone production, affecting testicular function (secondary hypogonadism) and spermatogenesis. This review provides an overview of the available evidence on the potential association between ICIs and the disruption of spermatogenesis, with special focus on ICIs targeting cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), programmed death protein 1 (PD-1) and programmed death-ligand 1 (PD-L1). Moreover, it highlights the need for further investigations and encourages the discussion of associated risks and fertility-preservation considerations between clinicians and patients.

20.
Methods Mol Biol ; 2770: 135-149, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38351452

RESUMEN

Testes have a complex architecture that is compartmentalized into seminiferous tubules with a diameter of approximatively 200 µm in which the germ cells differentiate, surrounded by a basement membrane and interstitium. 3D bioprinting might be used to recreate the compartmentalized testicular architecture in vitro. Directed by a software program, pneumatic microextrusion printers can deposit 3D layers of hydrogel-encapsulated interstitial cells in a controlled manner by applying pressure. Once macroporous-shaped scaffolds resembling seminiferous tubules have been bioprinted with interstitial cells, the epithelial cell fraction can be seeded in the macropores to resemble the in vivo testicular architecture. Moreover, macropores can serve as a delimitation for all testicular cells to reorganize and improve the supply of nutrients to cells through the 3D constructs.


Asunto(s)
Bioimpresión , Espermatogénesis , Masculino , Animales , Ratones , Testículo , Túbulos Seminíferos , Andamios del Tejido , Células Intersticiales del Testículo , Hidrogeles , Ingeniería de Tejidos , Impresión Tridimensional
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