RESUMO
Leydig cells are essential components of testicular interstitial tissue and serve as a primary source of androgen in males. A functional deficiency in Leydig cells often causes severe reproductive disorders; however, the transcriptional programs underlying the fate decisions and steroidogenesis of these cells have not been fully defined. In this study, we report that the homeodomain transcription factor PBX1 is a master regulator of Leydig cell differentiation and testosterone production in mice. PBX1 was highly expressed in Leydig cells and peritubular myoid cells in the adult testis. Conditional deletion of Pbx1 in Leydig cells caused spermatogenic defects and complete sterility. Histological examinations revealed that Pbx1 deletion impaired testicular structure and led to disorganization of the seminiferous tubules. Single-cell RNA-seq analysis revealed that loss of Pbx1 function affected the fate decisions of progenitor Leydig cells and altered the transcription of genes associated with testosterone synthesis in the adult testis. Pbx1 directly regulates the transcription of genes that play important roles in steroidogenesis (Prlr, Nr2f2 and Nedd4). Further analysis demonstrated that deletion of Pbx1 leads to a significant decrease in testosterone levels, accompanied by increases in pregnenolone, androstenedione and luteinizing hormone. Collectively, our data revealed that PBX1 is indispensable for maintaining Leydig cell function. These findings provide insights into testicular dysgenesis and the regulation of hormone secretion in Leydig cells.
Assuntos
Infertilidade Masculina , Células Intersticiais do Testículo , Fator de Transcrição 1 de Leucemia de Células Pré-B , Testículo , Testosterona , Animais , Masculino , Células Intersticiais do Testículo/metabolismo , Células Intersticiais do Testículo/patologia , Fator de Transcrição 1 de Leucemia de Células Pré-B/metabolismo , Fator de Transcrição 1 de Leucemia de Células Pré-B/genética , Camundongos , Testosterona/metabolismo , Testículo/metabolismo , Testículo/patologia , Infertilidade Masculina/genética , Infertilidade Masculina/patologia , Infertilidade Masculina/metabolismo , Diferenciação Celular/genética , Espermatogênese/genética , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
Klinefelter syndrome (KS), also known as 47, XXY, is characterized by a distinct set of physiological abnormalities, commonly including infertility. The molecular basis for Klinefelter-related infertility is still unclear, largely because of the cellular complexity of the testis and the intricate endocrine and paracrine signaling that regulates spermatogenesis. Here, we demonstrate an analysis framework for dissecting human testis pathology that uses comparative analysis of single-cell RNA-sequencing data from the biopsies of 12 human donors. By comparing donors from a range of ages and forms of infertility, we generate gene expression signatures that characterize normal testicular function and distinguish clinically distinct forms of male infertility. Unexpectedly, we identified a subpopulation of Sertoli cells within multiple individuals with KS that lack transcription from the XIST locus, and the consequence of this is increased X-linked gene expression compared to all other KS cell populations. By systematic assessment of known cell signaling pathways, we identify 72 pathways potentially active in testis, dozens of which appear upregulated in KS. Altogether our data support a model of pathogenic changes in interstitial cells cascading from loss of X inactivation in pubertal Sertoli cells and nominate dosage-sensitive factors secreted by Sertoli cells that may contribute to the process. Our findings demonstrate the value of comparative patient analysis in mapping genetic mechanisms of disease and identify an epigenetic phenomenon in KS Sertoli cells that may prove important for understanding causes of infertility and sex chromosome evolution.
Assuntos
Infertilidade Masculina/patologia , Síndrome de Klinefelter/complicações , Células Intersticiais do Testículo/patologia , Células de Sertoli/patologia , Análise de Célula Única/métodos , Testículo/patologia , Transcriptoma , Humanos , Infertilidade Masculina/etiologia , Infertilidade Masculina/metabolismo , Síndrome de Klinefelter/cirurgia , Células Intersticiais do Testículo/metabolismo , Masculino , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Células de Sertoli/metabolismo , Espermatogênese , Testículo/metabolismo , Inativação do Cromossomo XRESUMO
Anastomosing hemangioma is a recently described vascular neoplasm, initially identified in the male genitourinary tract. Since its first description, it has been reported at multiple anatomic sites, including rare cases in the female genital tract, most in the ovary. Herein we report the largest series to date of 12 ovarian anastomosing hemangiomas identified at our institution over a 15-yr period. The patients' age at the time of resection ranged from 50 to 76 yr (median: 62 yr), 3 patients presented with symptomatic pelvic masses, 3 tumors were identified by imaging studies, and the remaining 6 were incidentally discovered in ovaries removed for other indications. All tumors were unilateral, occurred at the ovarian hilum, and contained a vaguely lobulated architecture with sinusoidal-like vessels lined by hobnail endothelial cells with minimal to no cytologic atypia. A rim of luteinized/Leydig cells with abundant, eosinophilic cytoplasm and round, centrally placed nuclei surrounding the hemangioma was present in 9/12 tumors. Reinke crystals were observed in 3 of these 9 tumors. The volume of luteinized cells relative to the vascular proliferation ranged from 2% to 30%. All tumors with luteinized/Leydig cells also displayed numerous small eosinophilic, globular intracytoplasmic inclusions within the endothelial cells. The 3 tumors without luteinized/Leydig cells were exclusively intravascular lesions. Despite the frequent presence of luteinization/Leydig cells none of the patients experienced hormonal manifestations. Awareness of this rare benign ovarian entity is important, as its association with luteinized cells/Leydig cell hyperplasia (often exuberant) may be misinterpreted as a steroid cell tumor, Leydig-cell tumor, or as a mixed stromal-vascular tumor.
Assuntos
Hemangioma , Neoplasias Ovarianas , Neoplasias de Tecidos Moles , Humanos , Masculino , Feminino , Ovário/cirurgia , Ovário/patologia , Células Intersticiais do Testículo/patologia , Hiperplasia/patologia , Células Endoteliais/patologia , Hemangioma/cirurgia , Hemangioma/química , Hemangioma/patologia , Neoplasias Ovarianas/cirurgia , Neoplasias Ovarianas/patologia , Neoplasias de Tecidos Moles/patologiaRESUMO
Urogenital tract abnormalities are among the most common congenital defects in humans. Male urogenital development requires Hedgehog-GLI signaling and testicular hormones, but how these pathways interact is unclear. We found that Gli3XtJ mutant mice exhibit cryptorchidism and hypospadias due to local effects of GLI3 loss and systemic effects of testicular hormone deficiency. Fetal Leydig cells, the sole source of these hormones in developing testis, were reduced in numbers in Gli3XtJ testes, and their functional identity diminished over time. Androgen supplementation partially rescued testicular descent but not hypospadias in Gli3XtJ mutants, decoupling local effects of GLI3 loss from systemic effects of androgen insufficiency. Reintroduction of GLI3 activator (GLI3A) into Gli3XtJ testes restored expression of Hedgehog pathway and steroidogenic genes. Together, our results show a novel function for the activated form of GLI3 that translates Hedgehog signals to reinforce fetal Leydig cell identity and stimulate timely INSL3 and testosterone synthesis in the developing testis. In turn, exquisite timing and concentrations of testosterone are required to work alongside local GLI3 activity to control development of a functionally integrated male urogenital tract.
Assuntos
Criptorquidismo/genética , Regulação da Expressão Gênica no Desenvolvimento , Células Intersticiais do Testículo/patologia , Proteínas do Tecido Nervoso/metabolismo , Diferenciação Sexual/genética , Proteína Gli3 com Dedos de Zinco/metabolismo , Animais , Criptorquidismo/patologia , Modelos Animais de Doenças , Proteínas Hedgehog/metabolismo , Humanos , Insulina/metabolismo , Células Intersticiais do Testículo/metabolismo , Masculino , Camundongos , Camundongos Transgênicos , Mutação , Proteínas do Tecido Nervoso/genética , Proteínas/metabolismo , Transdução de Sinais/genética , Testosterona/metabolismo , Proteína Gli3 com Dedos de Zinco/genéticaRESUMO
In the mammalian testis, two distinct populations of Sertoli cells (SCs), the immature SCs (ISCs) and adult SCs (ASCs), play significant roles in regulating the development and function of Leydig cells. However, the effect of different SC types on the function of Leydig cells is poorly understood. Here, our study showed that miR-145-5p expression was significantly different in SCs at different stages, with the highest expression observed in ISCs. Exosomes mediate the transfer of miR-145-5p from ISCs to Leydig cells. Overexpression of miR-145-5p in Leydig cells significantly downregulated steroidogenic gene expression and inhibited testosterone synthesis. Additionally, miR-145-5p functioned by directly targeted steroidogenic factor-1 (Sf-1) and downregulated the expression of SF-1, which further downregulated the expression of steroidogenic genes, induced accumulation of lipid droplets, and eventually suppressed testosterone production. These findings demonstrate that SC-derived miR-145-5p plays a significant role in regulating the functions of Leydig cells and may therefore serve as a diagnostic biomarker for male hypogonadism developmental abnormalities during puberty.
Assuntos
Exossomos/metabolismo , Células Intersticiais do Testículo/metabolismo , MicroRNAs/genética , Células de Sertoli/metabolismo , Fator Esteroidogênico 1/antagonistas & inibidores , Esteroides/biossíntese , Testículo/metabolismo , Animais , Exossomos/genética , Células Intersticiais do Testículo/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células de Sertoli/patologia , Fator Esteroidogênico 1/genética , Fator Esteroidogênico 1/metabolismo , Testículo/patologiaRESUMO
Galectin 3 is a multifunctional lectin implicated in cellular proliferation, differentiation, adhesion, and apoptosis. This lectin is broadly expressed in testicular somatic cells and germ cells, and is upregulated during testicular development. Since the role of galectin 3 in testicular function remains elusive, we aimed to characterize the role of galectin 3 in testicular physiology. We found that galectin 3 transgenic mice (Lgals3-/-) exhibited significantly decreased testicular weight in adulthood compared to controls. The transgenic mice also exhibited a delay to the first wave of spermatogenesis, a decrease in the number of germ cells at postnatal day 5 (P5) and P15, and defective Sertoli cell maturation. Mechanistically, we found that Insulin-like-3 (a Leydig cell marker) and enzymes involved in steroid biosynthesis were significantly upregulated in adult Lgals3-/- testes. These observations were accompanied by increased serum testosterone levels. To determine the underlying causes of the testicular atrophy, we monitored cellular apoptosis. Indeed, adult Lgals3-/- testicular cells exhibited an elevated apoptosis rate that is likely driven by downregulated Bcl-2 and upregulated Bax and Bak expression, molecules responsible for live/death cell balance. Moreover, the percentage of testicular macrophages within CD45+ cells was decreased in Lgals3-/- mice. These data suggest that galectin 3 regulates spermatogenesis initiation and Sertoli cell maturation in part, by preventing germ cells from undergoing apoptosis and regulating testosterone biosynthesis. Going forward, understanding the role of galectin 3 in testicular physiology will add important insights into the factors governing the development of germ cells and steroidogenesis and delineate novel biomarkers of testicular function.
Assuntos
Apoptose , Galectina 3/fisiologia , Células Intersticiais do Testículo/patologia , Células de Sertoli/patologia , Espermatogênese , Espermatozoides/patologia , Animais , Hormônio Foliculoestimulante/metabolismo , Células Intersticiais do Testículo/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células de Sertoli/metabolismo , Espermatozoides/metabolismo , Testosterona/metabolismoRESUMO
We report a 27 -year-old male referred because of hypergonadotropic hypogonadism with low testosterone and azoospermia. At 23 years of age, he underwent an excision of a hypoechoic 0.7 cm nodule of the left testicle. The pathological diagnosis was a Leydig cell tumor. In the right testicle, there were three nodules at ultrasound, the biggest measuring 0.6 cm. Four years later, the nodules in the right testicle were still present and the larger nodule was excised. The biopsy showed tubules with only Sertoli cells in the perinodular zone. Diffuse and nodular hyperplasia of the Leydig cells was found in the interstitium. The pathological diagnosis was Sertoli syndrome with severe hyperplasia of the Leydig cells. With testosterone therapy, LH decreased, and the nodules disappeared. Thereafter, upon interrupting therapy, LH increased, and the nodules reappeared in two occasions. Resuming testosterone treatment, the nodules disappeared again, suggesting a Leydig cell hyperplasia dependent on chronic LH stimulation.
Assuntos
Hipogonadismo , Testosterona , Masculino , Humanos , Adulto , Testosterona/uso terapêutico , Testosterona/farmacologia , Células Intersticiais do Testículo/patologia , Hiperplasia/patologia , Hipogonadismo/tratamento farmacológico , Hipogonadismo/patologia , Células de Sertoli/patologiaRESUMO
Experimental autoimmune orchitis (EAO) is a useful model to study organ-specific autoimmunity and chronic testicular inflammation. This model reflects testicular pathological changes reported in immunological infertility in men. Progression of EAO in rodents is associated with a significantly increased percentage of testicular endothelial cells and interstitial testicular blood vessels, indicating an ongoing angiogenic process. Vascular endothelial growth factor A (VEGFA), the main regulator of physiological and pathological angiogenesis, can stimulate endothelial cell proliferation, chemotaxis and vascular permeability. The aim of this study was to explore the role of VEGFA in the pathogenesis of testicular inflammation. Our results found VEGFA expression in Leydig cells, endothelial cells and macrophages in testis of rats with autoimmune orchitis. VEGFA level was significantly higher in testicular fluid and serum of rats at the end of the immunization period, preceding testicular damage. VEGF receptor (VEGFR) 1 is expressed mainly in testicular endothelial cells, whereas VEGFR2 was detected in germ cells and vascular smooth muscle cells. Both receptors were expressed in testicular interstitial cells. VEGFR2 increased after the immunization period in the testicular interstitium and VEGFR1 was downregulated in EAO testis. In-vivo-specific VEGFA inhibition by Bevacizumab prevented the increase in blood vessel number and reduced EAO incidence and severity. Our results unveil relevance of VEGFA-VEGFR axis during orchitis development, suggesting that VEGFA might be an early marker of testicular inflammation and Bevacizumab a therapeutic tool for treatment of testicular inflammation associated with subfertility and infertility.
Assuntos
Doenças Autoimunes/patologia , Neovascularização Patológica , Testículo/irrigação sanguínea , Testículo/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Inibidores da Angiogênese/farmacologia , Animais , Doenças Autoimunes/imunologia , Doenças Autoimunes/metabolismo , Doenças Autoimunes/prevenção & controle , Bevacizumab/farmacologia , Modelos Animais de Doenças , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Células Intersticiais do Testículo/metabolismo , Células Intersticiais do Testículo/patologia , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Orquite/imunologia , Orquite/metabolismo , Orquite/prevenção & controle , Codorniz/embriologia , Ratos Wistar , Transdução de Sinais , Testículo/efeitos dos fármacos , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
Perfluorotetradecanoic acid (PFTeDA) is one of perfluoroalkyl substances widely found in the environment. PFTeDA may cause the dysfunction of male reproductive system. However, whether PFTeDA affects the regeneration of Leydig cells remains unclear. The objective of this study was to examine the effects of short-term exposure of PFTeDA on the late-stage maturation of Leydig cells. Fifty-four adult Sprague-Dawley male rats were daily gavaged with PFTeDA (0, 10, or 20 mg/kg body weight) for 10 days, and then were injected intraperitoneally with ethylene dimethane sulfonate (EDS, 75 mg/kg body weight/once) to ablate Leydig cells to induce their regeneration. On day 21 (early stage) and 56 (late stage) after EDS, hormone levels, gene expression, and protein levels were measured. PFTeDA did not affect the early stage of Leydig cell regeneration, because it had no effect on serum testosterone, luteinizing hormone, and follicle-stimulating hormone levels, Leydig cell number, and its gene and protein expression. PFTeDA significantly reduced serum testosterone level and down-regulated the expression of Leydig cell genes (Cyp11a1, Hsd3b1, Cyp17a1, Hsd17b3, Hsd11b1, and Insl3) and their proteins (CYP11A1, HSD3B1, CYP17A1, HSD17B3, and INSL3), decreased the phosphorylation of AKT1 and ERK1/2, as well as lowered sperm count in the epididymis at 20 mg/kg. In conclusion, short-term exposure to PFTeDA blocks the late-stage maturation of Leydig cells.
Assuntos
Proliferação de Células/efeitos dos fármacos , Fluorocarbonos/toxicidade , Células Intersticiais do Testículo/efeitos dos fármacos , Animais , Hormônio Foliculoestimulante/sangue , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Células Intersticiais do Testículo/enzimologia , Células Intersticiais do Testículo/patologia , Hormônio Luteinizante/sangue , Masculino , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais , Testosterona/sangue , Fatores de TempoRESUMO
Perfluoroundecanoic acid (PFUnA) is one of long-chain perfluoroalkyl carboxylic acids. However, the effect of PFUnA on pubertal development of Leydig cells remains unclear. The goal of this study was to investigate the effect of PFUnA on Leydig cell development in pubertal male rats. We orally dosed male Sprague-Dawley rats (age 35 days) with PFUnA at doses of 0, 1, 5, and 10 mg/kg/day from postnatal day (PND) 35 to PND 56. Serum testosterone and luteinizing hormone levels were remarkably reduced by PFUnA at ≥1 mg/kg while serum follicle-stimulating hormone levels were lowered at 5 and 10 mg/kg. PFUnA down-regulated the expression of Lhcgr, Scarb1, Star, Cyp11a1, Hsd3b1, Cyp17a1, Hsd17b3, Hsd11b1, Insl3, Nr5a1, Fshr, Dhh, Sod1, and Sod2 and their proteins in the testis and the expression of Lhb and Fshb in the pituitary. PFUnA reduced Leydig cell number at 5 and 10 mg/kg. PFUnA induced oxidative stress and increased autophagy. These may result from the inhibition of phosphorylation of mTOR, AKT1, AKT2, and ERK1/2 in the testis. In conclusion, PFUnA exhibits inhibitory effects on pubertal Leydig cell development possibly via inducing oxidative stress and increasing autophagy.
Assuntos
Autofagia/efeitos dos fármacos , Ácidos Graxos/toxicidade , Fluorocarbonos/toxicidade , Células Intersticiais do Testículo/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Fatores Etários , Animais , Proteínas Relacionadas à Autofagia/metabolismo , Hormônio Foliculoestimulante/sangue , Regulação Enzimológica da Expressão Gênica , Células Intersticiais do Testículo/metabolismo , Células Intersticiais do Testículo/patologia , Hormônio Luteinizante/sangue , Masculino , Hipófise/efeitos dos fármacos , Hipófise/metabolismo , Ratos Sprague-Dawley , Desenvolvimento Sexual , Transdução de Sinais , Contagem de Espermatozoides , Espermatozoides/efeitos dos fármacos , Espermatozoides/patologia , Testosterona/sangueRESUMO
Phthalates as plasticizers are widely used in many consumer products. Dipentyl phthalate (DPeP) is one of phthalates. However, there are currently few data on whether DPeP exposure affects rat Leydig cell development. In this study, we investigated the effects of in utero DPeP exposure on Leydig cell development in the testes of male newborn and adult rats. From gestational days 14 to 21, Sprague-Dawley pregnant rats were gavaged vehicle (corn oil, control) or DPeP (10, 50, 100, and 500 mg/kg body weight/day). Testosterone and the expression of Leydig cell genes and proteins in the testis at birth and at postnatal day 56 were examined. DPeP dose-dependently reduced serum testosterone levels of male offspring at birth and at postnatal day 56 at 100 and 500 mg/kg and lowered serum luteinizing hormone levels at adult males at ≥10 mg/kg when compared with the control. In addition, DPeP increased number of fetal Leydig cells by inducing their proliferation but down-regulated the expression of Lhcgr, Scarb1, Star, Cyp11a1, Hsd3b1, Cyp17a1, Hsd17b3, and Insl3 in fetal Leydig cells per se. DPeP reduced number of adult Leydig cells by inducing cell apoptosis and down-regulated the expression of Lhcgr and Star in adult Leydig cells at postnatal day 56. DPeP lowered SIRT1 and BCL2 levels in the testis of adult rats. In conclusion, DPeP adversely affects both fetal and adult Leydig cell development after in utero exposure.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Intersticiais do Testículo/efeitos dos fármacos , Ácidos Ftálicos/toxicidade , Plastificantes/toxicidade , Efeitos Tardios da Exposição Pré-Natal , Testículo/efeitos dos fármacos , Fatores Etários , Animais , Apoptose/efeitos dos fármacos , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Regulação Enzimológica da Expressão Gênica , Idade Gestacional , Células Intersticiais do Testículo/metabolismo , Células Intersticiais do Testículo/patologia , Hormônio Luteinizante/sangue , Masculino , Exposição Materna , Gravidez , Ratos Sprague-Dawley , Testículo/embriologia , Testículo/metabolismo , Testosterona/sangueRESUMO
Hydroxysteroid (17ß) dehydrogenase type 3 (HSD17B3) deficiency causes a disorder of sex development in humans, where affected males are born with female-appearing external genitalia, but are virilized during puberty. The hormonal disturbances observed in the Hsd17b3 knockout mice (HSD17B3KO), generated in the present study, mimic those found in patients with HSD17B3 mutations. Identical to affected humans, serum T in the adult HSD17B3KO mice was within the normal range, while a striking increase was detected in serum A-dione concentration. This resulted in a marked reduction of the serum T/A-dione ratio, a diagnostic hallmark for the patients with HSD17B3 deficiency. However, unlike humans, male HSD17B3KO mice were born with normally virilized phenotype, but presenting with delayed puberty. In contrast to the current belief, data from HSD17B3KO mice show that the circulating T largely originates from the testes, indicating a strong compensatory mechanism in the absence of HSD17B3. The lack of testicular malignancies in HSD17B3KO mice supports the view that testis tumors in human patients are due to associated cryptorchidism. The HSD17B3KO mice presented also with impaired Leydig cell maturation and signs of undermasculinization in adulthood. The identical hormonal disturbances between HSD17B3 deficient knockout mice and human patients make the current mouse model valuable for understanding the mechanism of the patient phenotypes, as well as endocrinopathies and compensatory steroidogenic mechanisms in HSD17B3 deficiency.
Assuntos
17-Hidroxiesteroide Desidrogenases/fisiologia , Hormônios Esteroides Gonadais/sangue , Infertilidade Masculina/patologia , Células Intersticiais do Testículo/patologia , Mutação , 17-Hidroxiesteroide Desidrogenases/deficiência , 17-Hidroxiesteroide Desidrogenases/genética , Animais , Feminino , Infertilidade Masculina/etiologia , Células Intersticiais do Testículo/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disease caused by a mutation in the IDUA gene, which codes α-L-iduronidase (IDUA), a lysosomal hydrolase that degrades two glycosaminoglycans (GAGs): heparan sulfate (HS) and dermatan sulfate (DS). GAGs are macromolecules found mainly in the extracellular matrix and have important signaling and structural roles which are essential to the maintenance of cell and tissue physiology. Nondegraded GAGs accumulate in various cell types, which characterizes MPS I as a multisystemic progressive disease. Many tissues and vital organs have been described in MPS I models, but there is a lack of studies focused on their effects on the reproductive tract. Our previous studies indicated lower sperm production and morphological damage in the epididymis and accessory glands in male MPS I mice, despite their ability to copulate and to impregnate females. Our aim was to improve the testicular characterization of the MPS I model, with a specific focus on ultrastructural observation of the different cell types that compose the seminiferous tubules and interstitium. We investigated the testicular morphology of 6-month-old male C57BL/6 wild-type (Idua+/+) and MPS I (Idua-/-) mice. We found vacuolated cells widely present in the interstitium and important signs of damage in myoid, Sertoli and Leydig cells. In the cytoplasmic region of Sertoli cells, we found an increased number of vesicles with substrates under digestion and a decreased number of electron-dense vesicles similar to lysosomes, suggesting an impaired flux of substrate degradation. Conclusions: Idua exerts an important role in the morphological maintenance of the seminiferous tubules and the testicular interstitium, which may influence the quality of spermatogenesis, having a greater effect with the progression of the disease.
Assuntos
Glicosaminoglicanos/genética , Doenças por Armazenamento dos Lisossomos/genética , Mucopolissacaridose I/genética , Células de Sertoli/patologia , Animais , Modelos Animais de Doenças , Progressão da Doença , Humanos , Células Intersticiais de Cajal/metabolismo , Células Intersticiais de Cajal/patologia , Células Intersticiais do Testículo/metabolismo , Células Intersticiais do Testículo/patologia , Doenças por Armazenamento dos Lisossomos/metabolismo , Doenças por Armazenamento dos Lisossomos/patologia , Masculino , Camundongos , Mucopolissacaridose I/metabolismo , Mucopolissacaridose I/patologia , Mutação/genética , Células de Sertoli/metabolismo , Espermatozoides/metabolismo , Espermatozoides/patologiaRESUMO
Humans are exposed to phthalates ubiquitously, which may threaten health. However, whether di-n-octyl phthalate can prevent pubertal sexual maturity is still elusive. In this study, male Sprague Dawley rats (age 35 days) were treated daily by gavage with 0, 10, 100, and 1000 mg/kg body weight of di-n-octyl phthalate from day 35 to day 49 after birth. Di-n-octyl phthalate significantly reduced serum testosterone levels at doses of 100 and 1000 mg/kg, but increased serum luteinizing hormone levels of 1000 mg/kg and decreased testosterone/luteinizing hormone ratio at ≥10 mg/kg, without affecting serum follicle-stimulating hormone levels. Di-n-octyl phthalate significantly induced Leydig cell hyperplasia (increased number of CYP11A1-positive Leydig cells) at 100 and 1000 mg/kg. Di-n-octyl phthalate down-regulates the gene expression of Cyp11a1, Hsd3b1 and Insl3 in individual Leydig cells. Di-n-octyl phthalate can also reduce the number of sperm in the epididymis. Di-n-octyl phthalate increased phosphorylated AKT1/AKT2 without affecting their total proteins, but increased the total protein and phosphorylated protein of ERK1/2 and GSK-3ß. Primary immature Leydig cells isolated from 35-day-old rats were treated with 0-50 µM di-n-octyl phthalate for 3 h. This phthalate inhibited androgen production under basal, LH-stimulated, and cAMP-stimulated conditions by 5 and 50 µM in vitro via down-regulating Cyp11a1 expression but up-regulating Srd5a1 expression in vitro. In conclusion, di-n-octyl phthalate induces hypergonadotropic hypogonadism caused by Leydig cell hyperplasia but reduced steroidogenic function and prevents sperm production.
Assuntos
Substâncias Perigosas/toxicidade , Hipogonadismo/induzido quimicamente , Ácidos Ftálicos/toxicidade , Androgênios/metabolismo , Animais , Regulação para Baixo , Glicogênio Sintase Quinase 3 beta/metabolismo , Hiperplasia/metabolismo , Hiperplasia/patologia , Hipogonadismo/metabolismo , Hipogonadismo/patologia , Células Intersticiais do Testículo/metabolismo , Células Intersticiais do Testículo/patologia , Hormônio Luteinizante/metabolismo , Masculino , Fosforilação , Ratos , Ratos Sprague-Dawley , Maturidade Sexual , Espermatozoides/metabolismo , Testículo/metabolismo , Testosterona/metabolismo , Regulação para CimaRESUMO
For decades, numerous chemical pollutants have been described to interfere with endogenous hormone metabolism/signaling altering reproductive functions. Among these endocrine disrupting substances, Bisphenol A (BPA), a widely used compound, is known to negatively impact germ and somatic cells in the testis. Physical agents, such as ionizing radiation, were also described to perturb spermatogenesis. Despite the fact that we are constantly exposed to numerous environmental chemical and physical compounds, very few studies explore the impact of combined exposure to chemical and physical pollutants on reproductive health. The aim of this study was to describe the impact of fetal co-exposure to BPA and IR on testicular function in mice. We exposed pregnant mice to 10 µM BPA (corresponding to 0.5 mg/kg/day) in drinking water from 10.5 dpc until birth, and we irradiated mice with 0.2 Gy (γ-ray, RAD) at 12.5 days post-conception. Co-exposure to BPA and γ-ray induces DNA damage in fetal germ cells in an additive manner, leading to a long-lasting decrease in germ cell abundance. We also observed significant alteration of adult steroidogenesis by RAD exposure independently of the BPA exposure. This is illustrated by the downregulation of steroidogenic genes and the decrease of the number of adult Leydig cells. As a consequence, courtship behavior is modified, and male ultrasonic vocalizations associated with courtship decreased. In conclusion, this study provides evidence for the importance of broadening the concept of endocrine disruptors to include physical agents, leading to a reevaluation of risk management and regulatory decisions.
Assuntos
Compostos Benzidrílicos/toxicidade , Raios gama/efeitos adversos , Células Intersticiais do Testículo/metabolismo , Fenóis/toxicidade , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Lesões Experimentais por Radiação/metabolismo , Animais , Feminino , Células HeLa , Humanos , Células Intersticiais do Testículo/patologia , Masculino , Camundongos , Gravidez , Efeitos Tardios da Exposição Pré-Natal/patologia , Lesões Experimentais por Radiação/patologiaRESUMO
The ClC-2 channel plays a critical role in maintaining ion homeostasis in the brain and the testis. Loss-of-function mutations in the ClC-2-encoding human CLCN2 gene are linked to the white matter disease leukodystrophy. Clcn2-deficient mice display neuronal myelin vacuolation and testicular degeneration. Leukodystrophy-causing ClC-2 mutant channels are associated with anomalous proteostasis manifesting enhanced endoplasmic reticulum (ER)-associated degradation. The molecular nature of the ER quality control system for ClC-2 protein remains elusive. In mouse testicular tissues and Leydig cells, we demonstrated that endogenous ClC-2 co-existed in the same protein complex with the molecular chaperones heat shock protein 90ß (Hsp90ß) and heat shock cognate protein (Hsc70), as well as the associated co-chaperones Hsp70/Hsp90 organizing protein (HOP), activator of Hsp90 ATPase homolog 1 (Aha1), and FK506-binding protein 8 (FKBP8). Further biochemical analyses revealed that the Hsp90ß-Hsc70 chaperone/co-chaperone system promoted mouse and human ClC-2 protein biogenesis. FKBP8 additionally facilitated membrane trafficking of ClC-2 channels. Interestingly, treatment with the Hsp90-targeting small molecule 17-allylamino-17-demethoxygeldanamycin (17-AAG) substantially boosted ClC-2 protein expression. Also, 17-AAG effectively increased both total and cell surface protein levels of leukodystrophy-causing loss-of-function ClC-2 mutant channels. Our findings highlight the therapeutic potential of 17-AAG in correcting anomalous ClC-2 proteostasis associated with leukodystrophy.
Assuntos
Encéfalo/metabolismo , Canais de Cloreto/genética , Células Intersticiais do Testículo/metabolismo , Neurônios/metabolismo , Doença de Pelizaeus-Merzbacher/genética , Proteostase/genética , Animais , Benzoquinonas/farmacologia , Encéfalo/efeitos dos fármacos , Encéfalo/patologia , Células CHO , Canais de Cloro CLC-2 , Canais de Cloreto/deficiência , Cricetulus , Modelos Animais de Doenças , Degradação Associada com o Retículo Endoplasmático/efeitos dos fármacos , Regulação da Expressão Gênica , Células HEK293 , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Lactamas Macrocíclicas/farmacologia , Células Intersticiais do Testículo/efeitos dos fármacos , Células Intersticiais do Testículo/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/patologia , Doença de Pelizaeus-Merzbacher/tratamento farmacológico , Doença de Pelizaeus-Merzbacher/metabolismo , Doença de Pelizaeus-Merzbacher/patologia , Isoformas de Proteínas/deficiência , Isoformas de Proteínas/genética , Transdução de Sinais , Proteínas de Ligação a Tacrolimo/genética , Proteínas de Ligação a Tacrolimo/metabolismoRESUMO
Stem cell transplantation is a candidate method for the treatment of Leydig cell dysfunction-related diseases. However, there are still many problems that limit its clinical application. Here, we report the establishment of CXCR4-SF1 bifunctional adipose-derived stem cells (CXCR4-SF1-ADSCs) and their reparative effect on Leydig cell dysfunction. CD29+ CD44+ CD34- CD45- ADSCs were isolated from adipose tissue and purified by fluorescence-activated cell sorting (FACS). Infection with lentiviruses carrying the CXCR4 and SF1 genes was applied to construct CXCR4-SF1-ADSCs. The CXCR4-SF1-ADSCs exhibited enhanced migration and had the ability to differentiate into Leydig-like cells in vitro. Furthermore, the bifunctional ADSCs were injected into BPA-mediated Leydig cell damage model mice via the tail vein. We found that the CXCR4-SF1-ADSCs were capable of homing to the injured testes, differentiating into Leydig-like cells and repairing the deficiency in reproductive function caused by Leydig cell dysfunction. Moreover, we investigated the mechanism underlying SF1-mediated differentiation and testosterone synthesis in Leydig cells, and the B-box and SPRY Domain Containing Protein (BSPRY) gene was proposed to be involved in this process. This study provides insight into the treatment of Leydig cell dysfunction-related diseases.
Assuntos
Células Intersticiais do Testículo/metabolismo , Proteínas/genética , Receptores CXCR4/genética , Transplante de Células-Tronco , Fator Esteroidogênico 1/genética , Adipócitos/metabolismo , Adipócitos/patologia , Animais , Diferenciação Celular/genética , Linhagem da Célula/genética , Células Cultivadas , Células Intersticiais do Testículo/patologia , Células Intersticiais do Testículo/transplante , Masculino , Células-Tronco Mesenquimais/metabolismo , Camundongos , Transdução de Sinais/genética , Testículo/metabolismo , Testículo/patologiaRESUMO
The desert hedgehog (Dhh) is crucial for spermatogenesis and Leydig cell differentiation, but little is known regarding its physiological function in cartilage. In this study, Dhh mRNA was abundant in antler chondrocytes, where it advanced cell proliferation concomitant with accelerated transition from the G1 to the S phase and induced elevation of the hypertrophic chondrocyte markers, Col X and Runx2. Silencing of Ptch1 resulted in appreciable Smo accumulation and enhanced rDhh stimulation of Smo, whose impediment by cyclopamine obscured the proliferative function of Dhh and alleviated its guidance of chondrocyte differentiation. Further analysis evidenced the noteworthy positive action of Smo in the bridging between Dhh and Gli transcription factors. Obstruction of Gli1 by GANT58 caused the failed stimulation of Col X and Runx2 by rDhh. Analogously, siRNA against Gli1-3 hindered chondrocyte differentiation in the context of rDhh. Simultaneously, Gli transcription factors mediated the regulation of Dhh on Foxa1, Foxa2, and Foxa3, whose knockdown impaired chondrocyte differentiation. Attenuation of Foxa antagonized the augmentation of Col X and Runx2 generated by rDhh. Collectively, Dhh signaling through its target Foxa appears to induce antler chondrocyte proliferation and differentiation.
Assuntos
Chifres de Veado/crescimento & desenvolvimento , Condrogênese/genética , Fatores de Transcrição Forkhead/genética , Espermatogênese/genética , Animais , Chifres de Veado/metabolismo , Cartilagem/crescimento & desenvolvimento , Cartilagem/metabolismo , Ciclo Celular/genética , Diferenciação Celular/genética , Condrócitos/metabolismo , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Cervos/genética , Cervos/crescimento & desenvolvimento , Proteínas Hedgehog/genética , Células Intersticiais do Testículo/citologia , Células Intersticiais do Testículo/patologia , Masculino , Transdução de SinaisRESUMO
Studies with 6-n-propyl-2-thiouracil (PTU) in laboratory rodents have shown that transient neonatal hypothyroidism leads to increased Sertoli cell (SC) number, testis size and sperm production. However, scarce and inconclusive data are available for farm animals. In the present study, Piau pigs received PTU in a gel capsule containing 8 mg/kg of body weight for 14 weeks starting from the first week of age, whereas control animals received only the vehicle. Blood samples were collected during the experimental period for hormonal evaluation in the serum. The animals were orchiectomized at adulthood and had their testes used for histomorphometric analysis. Indicating that the PTU concentration used was effective in promoting hypothyroidism, PTU-treated pigs showed a 30% lower body weight and reduced thyroxine levels (p < 0.05) during the treatment period. At adulthood, the body weight was similar in both groups but, surprisingly, PTU-treated pigs showed 30% lower testis weight (p < 0.05). In general, treated pigs presented increased follicle-stimulating hormone levels, whereas testosterone levels tended to be lower from 9 to 23 weeks of age. No significant differences were observed for estradiol, Leydig cell volume and number, tubular diameter, SC number per gram of testis, SC efficiency and meiotic index. However, seminiferous tubule occupancy, total tubular length, SC number per testis, and daily sperm production per testis and per gram of testis (DSP/g/T) were significantly lower (p < 0.05) in PTU-treated pigs. Therefore, in contrast to laboratory rodents, our results showed that SC proliferation and DSP/g/T (spermatogenic efficiency) in Piau pigs is diminished by postnatal PTU treatment.
Assuntos
Antimetabólitos/toxicidade , Hipotireoidismo/patologia , Propiltiouracila/toxicidade , Células de Sertoli/patologia , Espermatogênese/efeitos dos fármacos , Espermatozoides/patologia , Animais , Animais Recém-Nascidos , Contagem de Células , Hipotireoidismo/induzido quimicamente , Células Intersticiais do Testículo/efeitos dos fármacos , Células Intersticiais do Testículo/patologia , Masculino , Túbulos Seminíferos/efeitos dos fármacos , Túbulos Seminíferos/patologia , Células de Sertoli/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , SuínosRESUMO
Vinclozolin is a common dicarboximide fungicide used to protect crops from diseases. It is also an endocrine disruptor and is thought to be related to abnormalities of the reproductive tract. However, its mechanism of inducing abnormalities of the male reproductive tract is still unclear. The purpose of this study was to study the effect of gestational vinclozolin exposure on the development of rat fetal Leydig cells. Female pregnant Sprague-Dawley rats were exposed to vinclozolin (0, 25, 50, and 100 mg/kg body weight/day) by gavage from gestational day 14-21. Vinclozolin dose-dependently reduced serum testosterone levels at doses of 50 and 100 mg/kg and the anogenital distance at 100 mg/kg. RNA-seq, qPCR, and Western blotting showed that vinclozolin down-regulated the expression of Nr5a1, Sox9, Lhcgr, Cyp11a1, Hsd3b1, Hsd17b3, Amh, Pdgfa, and Dhh and their encoded proteins. Vinclozolin reduced the number of NR2F2-positive stem Leydig cells at a dose of 100 mg/kg and enhanced autophagy in the testes. In conclusion, vinclozolin disrupts reproductive tract development and testis development in male fetal rats via several pathways.