Efecto de la Vitamina E en Células Peritubulares de Testículos de Ratones Mus musculus Tratados con Ácido Valproico Durante el Estado Fetal y Pubertad / Effect of Vitamin E on Peritubular Cells from Testis of Mus musculus Mice Treated with Valproic Acid During the Fetal Stage and Puberty

El ácido valproico (VPA) es un fármaco antiepiléptico teratógenico que, al ser administrado durante etapas tempranas del embarazo, puede producir alteraciones en el desarrollo embriofetal, las que se manifiestan tanto a nivel del sistema nervioso como del testículo. No obstante, se ha reportado que la administración de vitamina E (VE) podría revertir dichas alteraciones. El objetivo del presente estudio fue determinar el efecto protector de la VE a nivel testicular en fetos y ratones púberes expuestos a VPA durante la fase embrionaria de su desarrollo. Se utilizó un total de 30 ratones hembra adultas gestantes (Mus musculus) cepa BALB/c, las cuales se dividieron en 6 grupos. El estudio contempló el análisis de fetos machos a los 17,5 días post-coital (dpc) y machos juveniles a las 6 semanas post-natal. A los grupos 1 y 4 se les administró 0,3 mL de solución fisiológica (grupos control para 17,5 dpc y 6 semanas postnatal, respectivamente). A los grupos 2 y 5 se les suministró la cantidad de 600 mg/kg de VPA (grupos VPA), en tanto que a los grupos 3 y 6 se les aplicó la misma dosis de VPA complementada con 200 UI de VE (grupos VPA+VE). Se describió la histología normal y patológica del compartimento peritubular del testículo. En los grupos VPA se evidenció una degeneración de la pared peritubular, y atrofia de túbulos seminíferos, así como exfoliación de las células germinales. Por el contrario, en los grupos VPA+VE tales signos no fueron observados y la morfología presentó aspecto normal solo con algunas alteraciones focales. Estos resultados corroboran el hecho que la administración de VE contrarresta en parte, los efectos deletéreos que ocasiona el VPA.

SUMMARY: Valproic acid (VPA) is a teratogenic antiepileptic drug that, when administered during the early stages of pregnancy, can produce alterations in embryo-fetal development, which manifest both at the level of the nervous system and the testicle. However, it has been reported that the administration of vitamin E (VE) could reverse these alterations. The study aimed to determine the protective effect of VE at the testicular level in fetuses and pubertal mice exposed to VPA during the embryonic phase of their development. 30 pregnant adult female mice (Mus musculus) BALB/c strain were used, which were divided into 6 groups. The study included the analysis of male fetuses at 17.5 days post-coital (dpc) and juvenile males at 6 weeks post-natal. Groups 1 and 4 were administered 0.3 mL of physiological solution. Groups 2 and 5 were given 600 mg/kg of VPA (VPA groups), while groups 3 and 6 were given the same dose of VPA supplemented with 200 IU of VE (VPA+VE). The normal and pathological histology of the peritubular compartment of the testis was described. In the VPA groups, degeneration of the peritubular wall, and atrophy of the seminiferous tubules, as well as exfoliation of the germ cells, were evident. On the contrary, in the VPA+VE groups such signs were not observed and the morphology presented a normal appearance with only some focal alterations. These results corroborate the fact that the administration of VE partially counteracts the deleterious effects caused by VPA.

Clinicopathological characteristics of Klinefelter syndrome: a testicular biopsy analysis of 87 cases / 中华病理学杂志

Objective: To investigate the clinicopathological characteristics of testicular biopsies from Klinefelter syndrome (KS) patients. Methods: The testicular biopsy specimens of 87 patients with KS (a total of 107 biopsy specimens) were collected from the Department of Pathology, Peking University Third Hospital, Beijing, China from January 2017 to July 2022. All patients were diagnosed as KS by peripheral blood karyotyping analysis. The testicular histopathologic features, testicular volume and hormone levels were evaluated retrospectively. The histopathologic analysis was used to assess the quantity and morphology of Leydig cells, the spermatogenic state of seminiferous tubules, the thickening of the basement membrane of seminiferous tubules and the changes of stroma. Results: Leydig cell proliferative nodules were seen in 95.3% (102/107) of KS testicular biopsy tissues. The eosinophilic inclusion bodies and lipofuscin in Leydig cells were found in 52.3% (56/107) and 57.9% (62/107) of specimens, respectively. The Sertoli cell only seminiferous tubules and the hyalinized tubules were found in 66.4% (71/107) and 76.6% (82/107) of the examined tissues, respectively. The tubules with complete spermatogenic arrest were found in 15.9% (17/107) of specimens, and 5.6% (6/107) of the specimens showed low spermatogenesis or incomplete spermatogenic arrest. In 85.0% (91/107) of the specimens, increased thick-walled small vessels with hyaline degeneration were identified. Conclusions: The most common features of KS testicular specimens are Leydig cell proliferative nodules, hyaline degeneration of seminiferous tubules and proliferation of thick-walled blood vessels. Testicular biopsy specimens of KS are rare. The pathologists can make a tentative diagnosis of KS based on the histological findings, combined with the ultrasound and laboratory results, which is helpful for further diagnosis and treatment of KS.

The effectiveness of insulin therapy in the functional and morphological recovery of leydig cells in rats submitted to Diabetes mellitus

The effects of systemic insulin administration at different concentrations on the testicular tissue of diabetic adult rats, induced by streptozotocin, are evaluated by the morphological analysis of spermatogenic process. Twenty-four adult male rats were divided into 1) Control Group: they received citrate buffer, by intraperitoneal injection; 2) Diabetic Group: induced by intraperitoneal injection of streptozotocin (60 mg. kg-1 of body weight); 3) Insulin 50%: induced diabetes treated with half of standard dosage of insulin; 4) Insulin 100%: induced diabetes treated with standard dose of insulin. After eight weeks, animals were weighted and anesthetized; testicles were removed and processed in resin. Body and testicular weight of diabetic rats decreased when compared to that of control. Parameters increased with insulin therapy. Testosterone levels were low in diabetic animals but rates recovered after insulin therapy. Nuclear diameter and volume of Leydig cells decreased in diabetic rats although they significantly increased after insulin therapy. Results showed that the administration of insulin in diabetic rats promoted a protective effect of testicular parenchyma, enhancing efficient recovery on testosterone levels and increase in daily sperm production.

Short-term exposure to gossypol causes reversible reproductive toxicity and nephrotoxicity in mice / 南方医科大学学报

OBJECTIVE@#To study the toxic effects of short-term exposure to gossypol on the testis and kidney in mice and whether these effects are reversible.@*METHODS@#Twenty 7 to 8-week-old male mice were randomized into blank control group, solvent control group, gossypol treatment group and drug withdrawal group. In the former 3 groups, the mice were subjected to daily intragastric administration of 0.3 mL of purified water, 1% sodium carboxymethylcellulose solution, and 30 mg/mL gossypol solution for 14 days, respectively; In the drug withdrawal group, the mice were treated with gossypol solution in the same manner for 14 days followed by treatment with purified water for another 14 days. After the last administration, the mice were euthanized and tissue samples were collected. The testicular tissue was weighed and observed microscopically with HE and PAS staining; the kidney tissue was stained with HE and examined for mitochondrial ATPase activity.@*RESULTS@#Compared with those in the control group, the mice with gossypol exposure showed reduced testicular seminiferous epithelial cells with rounded seminiferous tubules, enlarged space between the seminiferous tubules, interstitium atrophy of the testis, and incomplete differentiation of the spermatogonia. The gossypol-treated mice also presented with complete, non-elongated spermatids, a large number of cells in the state of round spermatids, and negativity for acrosome PAS reaction; diffuse renal mesangial cell hyperplasia, increased mesangial matrix, and adhesion of the mesangium to the wall of the renal capsule were observed, with significantly shrinkage or even absence of the lumens of the renal capsules and reduced kidney mitochondrial ATPase activity. Compared with the gossypol-treated mice, the mice in the drug withdrawal group showed obvious recovery of morphologies of the testis and the kidney, acrosome PAS reaction and mitochondrial ATPase activity.@*CONCLUSIONS@#Shortterm treatment with gossypol can cause reproductive toxicity and nephrotoxicity in mice, but these toxic effects can be reversed after drug withdrawal.

Gonadal differentiation during embryonic and fetal development of male New Zealand rabbits (Oryctolagus cuniculus) / Diferenciación gonadal durante el desarrollo embrionario y fetal de conejos machos neozelandeses (Oryctolagus cuniculus)

SUMMARY: The rabbit is considered an ideal animal model for studies that describe abnormalities in the testicles due to the similar morphogenetic mechanisms of sexual development and diseases commonly found in humans. The aim of this study was to determine the male sexual differentiation of the New Zealand rabbit (Oryctolagus cuniculus) through development. The gestational age was estimated and classified as 9, 12, 14, 16, 18, 20, 23 and 28 gestational days. The morphological and sexual determination were performed by histological analysis of the reproductive tract in the embryos and fetuses (9-28 days) as well as by immunohistochemistry- Desert hedgehog-Dhh- (testis-specific protein on Y chromosome- 16, 20, 23 days and adult rabbits). Gonads were observed from the 14th day in an undifferentiated stage and with homogeneous aspect. Sexual differentiation was observed from the 16th day with presence of cells forming gonadal cords and Dhh+ cells in the gonadal parenchyma. From the 18th gestational day testicular cords were observed, which evolved into organized seminiferous tubules. The formation of the efferent ducts and ductus deferens and epididymis was observed on the 20th and 23rd days, respectively. The differentiation of the external genitalia occurred from the 23rd days from the anogenital distance and was identified to identify the penile structures. In summary, the features of the sexual differentiation were determined by observation of the Dhh+ protein in embryos from the 16th day to adulthood, and the morphological particularities observed from the 18th gestational day, determined by differentiation of the external genitalia from the 23rd day.

RESUMEN: El conejo se considera un modelo animal ideal para estudios que describen anomalías a nivel testícular debido a que presenta mecanismos morfogenéticos similares al desa- rrollo sexual y enfermedades que se encuentran comúnmente en los seres humanos. El objetivo de este estudio fue determinar la diferenciación sexual masculina del conejo de Nueva Zelanda (Oryctolagus cuniculus) a través del desarrollo. La edad gestacional se estimó y clasificó en 9, 12, 14, 16, 18, 20, 23 y 28 días gestacionales. La determinación morfológica y sexual se realizó mediante análisis histológico del tracto reproductivo en los embriones y fetos (9 - 28 días) así como mediante inmunohistoquímica -Desert hedgehog-Dhh- (proteína testicular específica en el cromosoma Y- 16, 20, 23 días y conejos adultos). Las gónadas se observaron a partir del día 14 en un estadio indiferenciado y con aspecto homogéneo. Se observó diferenciación sexual a partir del día 16 con presencia de células formadoras de cordones gonadales y células Dhh+ en el parénquima gonadal. A partir del día 18 de gestación se observaron cordones testiculares, que evolucionaron a túbulos seminíferos organizados. La formación de los conductos eferentes, deferentes y del epidídimo se observó a los 20 y 23 días, respectivamente. La diferenciación de los genitales externos ocurrió a partir del día 23 desde la distancia anogenital y se utilizó para identificar las estructuras del pene. En conclusión, las características de la diferenciación sexual se determinaron mediante la observación de la proteína Dhh en embriones desde el día 16 hasta la edad adulta, y las particularidades morfológicas observadas a partir del día 18 de gestación, determinadas por diferenciación de los genitales externos a partir del día 23.

Effects of Corchorus olitorius and protacatechuic acid on diabetic rat testis tissue / Efectos de Corchorus olitorius y ácido protocatéquico en el tejido testicular de rata diabética

SUMMARY: The aim of this study is to investigate the effects of Protocatechuic acid and Corchorus olitorius on streptozotocin (STZ) induced diabetic rat testis tissue. Randomly selected Wistar Albino rats were divided into five groups as; Diabetes Mellitus, Diabetes Mellitus treated with Corchorus Olitorus (STZ+CO), Diabetes Mellitus treated with Protacatechuic acid (STZ+PCA), Corchorus olitorius (CO), Protocatechuic acid (PCA) and Control. Diabetic model was generated by intraperitoneal injection of 60 mg/kg Streptozotosin. After 48 hours of the STZ injection, blood samples were collected from tail vein in order to measure blood glycose levels. Over 250 mg/dL accepted as diabetic subjets and fed with 250 mg/kg Corchorus olitorius or 20 mg/kg PCA by oral gavage for three weeks. At the end of the experiment, right testes were removed and fixed in 10 % neutral formaldehyde for paraffine embedding. Sections were stained by HE, Masson trichrome, PAS and TUNEL for microscopic evaluation. Control, PCA-only and Corchorus olitorius-only treated group testes tissues showed a normal tissue organization, when degeneration in seminiferous tubules, the vacuolization, seperations in spermatogenic cell series, outpouring of cell groups in the lumen, vesicular body formation, liquid accumulation in the interstitial region and edema were observed in STZ induced diabetic models and untreated groups. Besides, higher amount of TUNEL (+) stained cells were determined in STZ group. On the other hand, blood glucose level and number of TUNEL (+) stained cells were decreased as a result of PCA and Corchorus olitorius treatment. Because of the reduction of blood glucose level and apoptotic cell numbers, PCA and Corchorus olitorius decreace the complications of diabetes mellitus induced rat testis.

RESUMEN: El objetivo de este estudio fue investigar los efectos del ácido protocatéquico y Corchorus olitorius sobre el tejido testicular de rata diabética inducida por estreptozotocina (STZ). Las ratas Wistar Albino fueron seleccionadas al azar y se dividieron en cinco grupos; Diabetes Mellitus, Diabetes Mellitus tratada con Corchorus olitorius (STZ + CO), Diabetes Mellitus tratada con ácido protocatéquico (STZ + PCA), Corchorus olitorius (CO), ácido protocatéquico (PCA) y Control. El modelo diabético se generó por inyección intraperitoneal de 60 mg/kg de estreptozotosina. Después de 48 horas de la inyección de STZ, se recogieron muestras de sangre de la vena de la cola para medir los niveles de glucosa. Niveles mayores a 250 mg/dL fueron considerados como especímenes diabéticos y alimentados con Corchorus olitorius de 250 mg/kg o PCA de 20 mg/kg por sonda oral durante tres semanas. Al final del experimento, se extirparon los testículos derechos y se fijaron en formaldehído neutro al 10 % para la inclusión en parafina. Las secciones se tiñeron con HE, tricromo de Masson, PAS y TUNEL para evaluación microscópica. Los tejidos de los testículos de los grupos control, tratados solo con PCA y con Corchorus olitorius mostraron una organización tisular normal. En cambio en modelos diabéticos inducidos por STZ y grupos no tratados se observó degeneración en los túbulos seminíferos, vacuolización, separaciones en series de células espermatogénicas, efusión de grupos celulares en la luz, formación del cuerpo vesicular, acumulación de líquido en la región intersticial y edema. Además, se determinó una mayor cantidad de células teñidas con TUNEL (+) en el grupo STZ. Por otro lado, el nivel de glucosa en sangre y el número de células teñidas con TUNEL (+) disminuyeron como resultado del tratamiento con PCA y Corchorus olitorius. Debido a la reducción del nivel de glucosa en sangre y el número de células apoptóticas, se observó que PCA y Corchorus olitorius disminuyen las complicaciones de los testículos de rata inducidos por diabetes mellitus.

Testicular features of semi-confined Nellore bullssubjected to immunocastration / Características testiculares de touros Nelore semiconfinadossubmetidos a imunocastração

This study aimed to evaluate the effect of immunocastration on the scrotal circumference, as well as in the macroscopic and microscopic testicular features in Nellore bulls bred under a semi-confining system for 91 days. Sixty animals were divided into two groups: 30 intact animals (non-vaccinated) and 30 immunocastrated animals. The immunocastrated animals were treated with two anti-GnRH vaccine doses (BoprivaTM - Zoetis Australia Pty Ltd). The scrotal circumference (SC) was measured on days 0 and 56. Testicular parameters: biometry traits (length, width, and height ­ cm; volume ­ cm3) and weight (g) was measured on day 91. Three testicles from each group were histologically processed to find the seminiferous tubule diameter and epithelium thickness (µm). Data were analyzed in the Minitab® 19 statistical software. The macroscopic features (SC, and testes weight, volume, length, width, and height) and the microscopic features (seminiferous tubule diameter and epithelium thickness) were compared between the two groups through Student's t-test and Mann-Whitney test. All the values were lower (p<0.05) in animals immunized against GnRH, except for the right testis width and seminiferous tubule diameter. Results indicate that immunization against GnRH affected testicular development.(AU)

O objetivo do presente estudo foi avaliar o efeito da imunocastração no perímetro escrotal, assim como nas características testiculares macroscópicas e microscópicas de touros Nelore criados em um sistema de semiconfinamento por 91 dias. Sessenta animais foram divididos em dois grupos: 30 animais inteiros (não vacinados) e 30 imunocastrados. Os animais imunocastrados receberam duas doses de uma vacina anti-GnRH (Bopriva® - Zoetis Ltda). O perímetro escrotal (PE) foi mensurado nos dias 0 e 56. Parâmetros testiculares: atributos biométricos (comprimento, largura e altura ­ cm; volume cm3) e peso (g) foram medidos no dia 91. Três testículos de cada grupo foram processados para histologia clássica a fim de avaliar o diâmetro e a espessura do epitélio dos túbulos seminíferos (µm). Os dados foram analisados pelo software estatístico Minitab® 19. As médias das características macroscópicas (PE, peso, volume, comprimento, largura e altura dos testículos) e das microscópicas (diâmetro e espessura do epitélio dos túbulos seminíferos) foram comparadas entre os dois grupos pelos testes t de Student e Mann-Whitney. Todos os valores foram menores (p<0,05) em animais imunizados contra o GnRH, exceto a altura média do testículo direito e o diâmetro do túbulo seminífero. Os resultados indicam que a imunização contra o GnRH afeta o desenvolvimento testicular.(AU)

Effects of ketoprofen in spermatogenesis of mice exposed to continuos hypobaric hypoxia / Efecto del ketoprofeno en la espermatogénesis de ratones expuestos a hipoxia hipobárica continua

Hypoxia hypobaric (HH) can cause alterations at testicular level, with temperature increase, intrascrotal alteration and deterioration of spermatogenesis. Nonsteroidal anti-inflammatory drugs (NSAIDs) such as ketoprofen have anti-angiogenic properties, and can decrease testicular abnormalities. The objective of the study was to evaluate the effect of ketoprofen on spermatogenesis of mice exposed to continuous hypobaric hypoxia. 78 Mus musculus CF-1 male mice 3 to 4 months old were used and subjected to HH in chamber at 4200 m. They were divided into 13 groups (G) of 6 animals: 10 with HH cycles (1, 2, 3, 4 and 8, lasting 8.3 days each cycle, two groups each) and 3 in normoxia (Nx). Intraperitoneal ketoprofen 25 mg/kg was administered every 4 days. Euthanasia of these animals was performed at the end of each cycle and in the case the Nx groups at the end of cycles 1, 4 and 8. Percentage of microhematocrit and reticulocytes were measured in blood smears and a morphometric and histopathological analysis of the height of the epithelium, the tubular diameter and the diameter of the tubular lumen was made. It was shown that hematocrit increases continuously up to 8 cycles, while reticulocytes increase up to 3 cycles. Continuous HH decreases the tubular diameter in a sustained manner and proportional to HH cycles, and the height increased only in the groups subjected to 8 cycles. The groups treated with ketoprofen saw a decrease in angiogenesis, presenting some degree of protection at the testicular level.

La hipoxia hipobárica (HH) puede provocar alteraciones a nivel testicular, con aumento de la temperatura, alteración intraescrotal y deterioro de la espermatogénesis. Los antiinflamatorios no esteroidales (AINEs) como el ketoprofeno tienen propiedades antiangiogénicas, pudiendo disminuir las alteraciones testiculares. El objetivo de estudio fue evaluar el efecto del ketoprofeno en la espermatogénesis de ratones expuestos a hipoxia hipobárica continua. Se utilizaron 78 ratones macho Mus musculus CF-1 de 3 a 4 meses de edad y se sometieron a HH en cámara a 4200 m. Se dividieron en 13 grupos (G) de 6 animales: 10 con ciclos de HH (1, 2, 3, 4 y 8, con duración de 8,3 días cada ciclo, dos grupos cada uno) y 3 en normoxia (Nx). Se administró ketoprofeno intraperitoneal 25 mg/kg cada 4 días. La eutanasia de estos animales se realizó al final de cada ciclo y en el caso los grupos Nx al final de los ciclos 1, 4 y 8. Se midió porcentaje de microhematocrito y reticulocitos en frotis de sangre y se hizo un análisis morfométrico e histopatológico de la altura del epitelio, el diámetro tubular y el diámetro de la luz tubular. Se evidenció que el hematocrito aumenta de manera continua hasta los 8 ciclos, en cambio los reticulocitos aumentan hasta los 3 ciclos. La HH continua disminuye el diámetro tubular de forma sostenida y proporcional a los ciclos de HH, y la altura aumentó sólo en los grupos sometidos a 8 ciclos. Los grupos tratados con ketoprofeno se vio una disminución de la angiogénesis, presentando algún grado de protección a nivel testicular.

Effect of melatonin on the onset of puberty in male juvenile rats / 대한해부학회지

Melatonin or N-acetyl-5-methoxytryptamine, the fascinating molecule secreted by the pineal gland. Melatonin has a close interaction with hypothalamic-pituitary-gonadal axis. In non-seasonal breeders like rat its exact role in reproduction is controvertible. So it is worth to explore the possible role of melatonin on the onset of puberty in male albino rats. Two groups of male rats aged 5 and 10 days were used for the study. In each group, there were three subgroups, each receiving melatonin for 5 days, 10 days or till the day of descent of testes. Similar subgroups were used as controls. Without handling, animals were observed daily for the onset of puberty. On the day of descent of testes, body weight of the animal was noted, blood was collected, serum was separated and used for radio immunoassay. For histomorphometric analysis, all morphometric measurements were done using an occular micrometer. Volume fraction of seminiferous tubules, intertubular connective tissue of testes, cortex and medulla of thymus were estimated by point count method. In both the age groups melatonin advanced the age on descent of testes, increased the body weight, organ weight. It also increased the serum hormone levels. So, in conclusion this study indicates that exogenous melatonin advances the onset of puberty in male albino wistar rats and this effect is more pronounced in the younger animals.

Histology of the testis and the epididymal ducts from hoary fox Lycalopex vetulus (Lund, 1842) / Histologia dos testículos e ductos epididimários da raposa-do-campo Lycalopex vetulus (Lund, 1842)

The hoary fox Lycalopex vetulus (Lund, 1842) is a small canid, endemic to Brazil, belonging to the Canidae family, widely distributed in the country, occurring records in different regions and habitats. The objective of this study is to describe morphologically the testicles and epididymal ducts of hoary fox. The animals, coming from the zoo of Federal University of Mato Grosso, Brazil, had died by natural causes. The male reproductive system was dissected to collect the testicles. The samples were fragmented and histologically examined. Microscopically, the testes were coated by the vaginal and albuginea tunic, formed by modeled dense connective tissue with large amount of collagen fibers. Into the organ, convoluted seminiferous tubules were surrounded by a basement membrane characterized by the presence of myoid and Sertoli cells and germinative epithelium composed by Between the seminiferous tubules, interstitial tissue composed of connective tissue, blood and lymph vessels and Leydig cells in polyhedral shape was present. The epididymal ducts showed pseudostratified columnar epithelium with secretory cells, in which stereocilia located on a basement membrane filled by myoid cells were found. The structures observed by us are very similar to those described for other mammals.(AU)

A raposa-do-campo Lycalopex vetulus (Lund, 1842) é um canídeo de pequeno porte, endêmico do Brasil, pertencente a família Canidae, com ampla distribuição no país, ocorrendo registros em várias regiões e habitats diferentes. Com base nessa informação, o objetivo deste trabalho é caracterizar morfologicamente os testículos e ductos epididimários da raposa-do-campo. O animal, oriundo do zoológico da Universidade Federal de Mato Grosso, Brasil, veio a óbito por causas naturais e o sistema reprodutor masculino foi dissecado para coleta dos testículos. As amostras retiradas foram fragmentadas e histológicamente examinadas. A partir das análises microscópicas dos testículos foram identificados: a túnica vaginal e albugínea, formada por tecido conjuntivo denso modelado, com grande quantidade de fibras colágenas; túbulos seminíferos enovelados e revestidos por epitélio germinativo e células de Sertoli, envolvidos por uma membrana basal com presença de células mioides; tecido intersticial entre os túbulos seminíferos constituído de tecido conjuntivo, vasos sanguíneos e linfáticos, e células de Leydig em formato poliédrico. Os ductos epididimários apresentaram epitélio cilíndrico pseudoestratificado com células secretoras dos quais projetam estereocílios, situados sobre uma membrana basal repleta por células mióides. As estruturas por nós observadas possuem muita semelhança com as demais descrições para mamíferos.(AU)

Stereological and morphological analysis of the effects of aging on spermatogenesis in rat testis

Abstract Purpose: To quantify, through stereological and morphometric analysis, spermatogenesis in rats undergoing the natural aging process. Methods: Seventy-two male Wistar rats were divided into 6 equal groups according to age at the time of killing: 3, 6, 9, 12, 18, and 24 months. All the rats were subjected orchiectomy and collection of testicular parenchymal fragments for histological and morphometric analysis. The numerical density of spermatids was calculated using a stereological study, and morphometric analysis was conducted to measure the height of the germinal epithelium and the area of the seminiferous tubules. Results: We found that the 18 and 24 months groups showed a significant reduction in the number of round spermatids. However, the height of the germinal epithelium was not significantly different between the groups. The area of seminiferous tubules was also significantly reduced in the elderly rats compared to that in the young ones. Conclusion: Aging of rats showed a significant reduction in the number of round spermatids and the area of the seminiferous tubules, more pronounced in the rats at 18 and 24 months of life.

Expression of CLAUDIN-11 in the testicular tissue of the patient with non-obstructive azoospermia and its clinical significance / 中华男科学杂志

<p><b>Objective</b>To study the expression of CLAUDIN-11 in the testis tissue of non-obstructive azoospermia (NOA) patients with different severities and investigate its clinical significance.</p><p><b>METHODS</b>Sixty-two NOA patients were divided into a hypospermatogenesis (HS) group (n = 30) and a Sertoli cell only syndrome (SCO) group (n =32). The expression of CLAUDIN-11 in the testicular tissue of the patients was detected by immunohistochemistry, that of CLAUDIN-11 mRNA determined by real-time fluorescence quantitative polymerase chain reaction (RT-qPCR), and the levels of serum reproductive hormones measured by chemiluminescent immunoassay.</p><p><b>RESULTS</b>Immunohistochemistry showed that the expression of CLAUDIN-11 was mainly in the cytoplasm of the Sertoli cells around the seminiferous tubule wall in the HS group, but diffusely distributed in the membrane of the Sertoli cells in the SCO group. RT-qPCR revealed a significantly lower expression of CLAUDIN-11 mRNA in the HS than in the SCO group (0.008 ± 0.001 vs 0.013 ± 0.002, t = 10.616, P<0.01). The level of serum luteotropic hormone (LH) was also markedly lower in the HS than in the SCO group (［3.62 ± 1.34］ vs ［4.96 ± 3.10］ IU/L, P<0.05) and so was that of follicle-stimulating hormone (FSH) (［5.36 ± 2.80］ vs ［10.65 ± 9.18］ IU/L, P<0.05).</p><p><b>CONCLUSIONS</b>The up-regulated expression of CLAUDIN-11 in Sertoli cells may play an important role in the development and progression of spermatogenic dysfunction in NOA patients.</p>

Lipoic acid protects spermatogenesis in male rats with ornidazole-induced oligoasthenozoospermia / 中华男科学杂志

<p><b>Objective</b>To study the protective effect of lipoic acid (LA) on the spermatogenic function of the male rats with oligoasthenozoospermia induced by ornidazole (ORN).</p><p><b>METHODS</b>Seventy male SD rats were equally randomized into groups A (solvent control: 1 ml 0.5% CMC-Na + 1 ml olive oil), B (low-dose ORN model: 400 mg/kg ORN suspension + 1 ml olive oil), C (low-dose ORN + low-dose LA treatment: 400 mg/kg ORN + 50 mg/kg LA), D (low-dose ORN + high-dose LA treatment: 400 mg/kg ORN + 100 mg/kg LA), E (high-dose ORN model: 800 mg/kg ORN suspension + 1 ml olive oil), F (high-dose ORN + low-dose LA treatment: 800 mg/kg ORN + 50 mg/kg LA), and G (high-dose ORN + high-dose LA treatment: 800 mg/kg ORN + 100 mg/kg LA), and treated respectively for 20 successive days. Then all the rats were sacrificed and the weights of the body, testis, epididymis and seminal vesicle obtained, followed by calculation of the organ index, determination of epididymal sperm concentration and motility, and observation of the histomorphological changes in the testis and epididymis by HE staining.</p><p><b>RESULTS</b>Compared with group A, group E showed significantly decreased body weight (［117.67 ± 11.53］ vs ［88.11 ± 12.65］ g, P < 0.01) and indexes of the testis (［1.06 ± 0.12］ vs ［0.65 ± 0.13］ %, P < 0.01) and epididymis (［0.21 ± 0.03］ vs ［0.17 ± 0.01］ %, P < 0.01). In comparison with group E, group F exhibited remarkable increases in the epididymal index (［0.17 ± 0.01］ vs ［0.20 ± 0.02］ %, P < 0.01), and so did group G in the body weight (［88.11 ± 12.65］ vs ［102.70 ± 16.10］ g, P < 0.05) and the indexes of the testis (［0.65 ± 0.13］ vs ［0.95 ± 0.06］ %, P < 0.01) and epididymis (［0.17 ± 0.01］ vs ［0.19 ± 0.02］ %, P < 0.05), but no obvious difference was observed in the index of seminal vesicle among different groups. Compared with group A, group B manifested significant decreases in sperm motility (［74.12 ± 8.73］ vs ［40.25 ± 6.08］ %, P < 0.01), and so did group E in sperm count (［38.59 ± 6.40］ vs ［18.67 ± 4.59］ ×105/100 mg, P < 0.01) and sperm motility (［74.12 ± 8.73］ vs ［27.58 ± 8.43］ %, P < 0.01). Sperm motility was significantly lower in group B than in C and D (［40.25 ± 6.08］ vs ［58.13 ± 7.62］ and ［76.04 ± 8.44］%, P < 0.01), and so were sperm count and motility in group E than in F and G (［18.67 ± 4.59］ vs ［25.63 ± 9.66］ and ［29.92 ± 4.15］ ×105/100 mg, P < 0.05 and P < 0.01; ［27.58 ± 8.43］ vs ［36.56 ± 11.08］ and ［45.05 ± 9.59］ %, P < 0.05 and P < 0.01). There were no obvious changes in the histomorphology of the testis and epididymis in groups A, B, C and D. Compared with group A, group E showed necrotic and exfoliated spermatogenic cells with unclear layers and disorderly arrangement in the seminiferous tubules and remarkably reduced sperm count with lots of noncellular components in the epididymal cavity, while groups F and G exhibited increased sperm count in the seminiferous tubules and epididymis lumen, also with exfoliation, unclear layers and disorderly arrangement of spermatogenic cells, but significantly better than in group E.</p><p><b>CONCLUSIONS</b>LA can reduce ORN-induced damage to the spermatogenetic function of rats, improve sperm quality, and protect the reproductive system.</p>

Ameliorative effect of vitamin B12 on seminiferous epithelium of cimetidine-treated rats: a histopathological, immunohistochemical and ultrastructural study / 대한해부학회지

Cimetidine is an H2 receptor antagonist that has an antiandrogenic effect. It intervenes with the conversion of testosterone into estrogen in the Sertoli cells with accompanying testicular structural changes. In the present study, the microscopic and the ultrastructural changes induced by cimetidine and the effect of vitamin B12 as a protective agent on rat testes were studied. Immunoexpression of estrogen receptor β (ERβ) in testes was evaluated. Twenty-four adult male rats were divided into four groups: control, cimetidine-treated, vitamin B12 treated, and combined cimetidine and vitamin B12 treated. The experimental rats were administered with cimetidine and/or vitamin B12 for 52 days. Group II rats showed marked atrophy of the seminiferous tubules with a significant increase in tubular diameter and decrease in the tubular luminal and epithelial areas. Ultrastructure of this group showed irregular Sertoli cells with basal cytoplasmic vacuolation and significantly thickened basement membrane. ERβ immunoexpression was similar to controls. Group III rats showed near normal seminiferous tubular structures with minimal cellular alterations and the immunoreactivity of the testicular sections was very close to normal. However, group IV rats showed markedly immunopositive detached cells, spermatids, and primary spermatocytes. Cimetidine interferes with the control of spermatogenesis as evidenced by microscopic and ultrastructural studies and affection of ERβ receptors and vitamin B12 has a protective action against this harmful effect.

Successful onco-testicular sperm extraction from a testicular cancer patient with a single testis and azoospermia / 대한생식의학회지

Onco-testicular sperm extraction is used to preserve fertility in patients with bilateral testicular tumors and azoospermia. We report the case of a testicular tumor in the solitary testis of a patient who had previously undergone successful contralateral orchiectomy and whose sperm was preserved by onco-testicular sperm extraction. A 35-year-old patient presented with swelling of his right scrotum that had lasted for 1 month. His medical history included a contralateral orchiectomy during childhood. Ultrasonography revealed a mosaic echoic area in his scrotum, suggesting a testicular tumor. The lesion was palpated within the normal testicular tissue along its edge and semen analysis showed azoospermia. Radical inguinal orchiectomy and onco-testicular sperm extraction were performed simultaneously. Motile spermatozoa were extracted from normal seminiferous tubules under microscopy and were frozen. Eventual intracytoplasmic sperm injection using the frozen spermatozoa is planned. Onco-testicular sperm extraction is an important fertility preservation method in patients with bilateral testicular tumors or a history of a previous contralateral orchiectomy.

An infertile patient with Y chromosome b1/b3 deletion presenting with congenital bilateral absence of the vas deferens with normal spermatogenesis / 대한생식의학회지

We report the case of a 46-year-old Chinese male patient who visited our clinic complaining of infertility. Semen analysis revealed azoospermia, and azoospermia factor c region partial deletion (b1/b3) was detected using Y chromosome microdeletion analysis. Testicular sperm extraction was performed after genetic counseling. The bilateral ductus deferens and a portion of the epididymis were absent, whereas the remaining epididymis was expanded. Motile intratesticular spermatozoa were successfully extracted from the seminiferous tubule. On histopathology, nearly complete spermatogenesis was confirmed in almost every seminiferous tubule. To our knowledge, this is the first case report of b1/b3 deletion with a congenital bilateral absence of the vas deferens and almost normal spermatogenesis.

Influence of prolonged flaxseed ( Linum usitatissimum ) consumption over epididymis and testicle histoarchitecture of Wistar rats / Influência do consumo prolongado de semente de linhaça ( Linum usitatissimum ) sobre a histoarquitetura dos testículo e epidídimo de ratos Wistar

Flaxseed is considered a functional food with several health benefits. However, because of its high phytoestrogen content, flaxseed influences hormone metabolism and affects the gonadal biomorphology. In this study, computerized histomorphometry was used to evaluate seminiferous and epididymal tubules, considering the different regions of the epididymis (head, body and tail) of rats subjected to a prolonged diet of flaxseed. Young adult male Wistar rats (n=20) were divided into 2 groups during their lactation period: Control Group (CG), fed casein-based meals and Flaxseed Group (FG), fed a 25% flaxseed meal. After 250 days of continuous ingestion, the animals were euthanized and a blood sample was collected. The testicles and epididymis were removed and fixed in buffered formalin solution. The samples were subjected to routine histological paraffin techniques and stained with hematoxilin and eosin. Immunostaining was performed using an antivimentin antibody for Sertoli cell identification. For morphometry, images of the slides were scanned and analyzed using Image J to determine the epithelial height, tubular and luminal diameter and tubular and luminal area. In the hormonal evaluation, FG had a higher serum concentration of estrogen (P=0.001), but no change was observed in the concentration of testosterone. The morphometric assay of seminiferous tubules and epididymal regions revealed no significant differences between the analyzed groups. Similarly, Sertoli cell quantification showed no significant differences in the FG (P=0.98). These results revealed that the continuous and prolonged intake of 25% flaxseed meals from gestation to 250 days of age, even with a significant increase in serum levels of estradiol, does not exert adverse effects on the testicular and epididymal structure or on the cells participating in the spermatogenesis of rats.(AU)

A semente de linhaça é considerada um alimento funcional com vários efeitos benéficos à saúde. Entretanto, devido ao seu elevado teor de fitoestrógenos, esta semente pode influenciar no metabolismo hormonal e interferir na biomorfologia gonadal. Neste estudo, utilizamos a histomorfometria computadorizada para avaliar os túbulos seminíferos e epididimários, considerando as diferentes regiões do epidídimo (cabeça, corpo e cauda) de ratos submetidos a uma dieta prolongada de semente de linhaça. Foram utilizados ratos Wistar machos adultos jovens (n=20) divididos em 2 grupos, durante o período de lactação: Grupo Controle (GC) a base de caseína e Grupo Linhaça (GL) alimentados com 25% de semente de linhaça. Ao final de 250 dias de ingestão contínua, os animais foram sacrificados e amostra de sangue foi coletada. Os testículos e epidídimos foram retirados e fixados em formol tamponado. As amostras foram submetidas ao processamento histológico de rotina para parafina e coradas em hematoxilina e eosina. Foi feita a imunomarcação com anticorpo antivimentina para identificação das células de Sertoli. Para morfometria, as imagens das lâminas foram digitalizadas e analisadas pelo software ImageJ para obtenção dos dados de altura epitelial, diâmetro e área tubular e luminal. Na avaliação hormonal o GL teve maior concentração de estrógeno sérico (p=0,001), mas nenhuma mudança na concentração de testosterona foi observada. Nos parâmetros morfométricos dos túbulos seminíferos e das regiões epididimárias, não houve diferenças significativas entre os grupos analisados. Da mesma forma, a quantificação das células de Sertoli não apresentaram diferenças significativas no GL (p=0,98). Estes resultados mostraram que o consumo contínuo e prolongado de 25% de semente de linhaça desde período gestacional até 250 dias de idade, mesmo com o aumento significativo nos níveis séricos de estradiol, não exerceram efeitos adversos sobre a estrutura testicular e epididimária, assim como nas células participantes da espermatogênese em ratos.(AU)

Short-term buserelin administration induces apoptosis and morphological changes in adult rat testes

Abstract Purpose: To investigate the effect of buserelin on gonadal structure and function in adult male rats. Methods: Twenty-four adult Wistar male rats were divided into three groups: two treated groups and controls. The first and second treated groups received 300 (low dose) and 500 (high dose) µg/kg buserelin, respectively, and the control group received normal saline. All groups were treated subcutaneously for five days. Results: The seminiferous tubular epithelial thickness was significant decreased in the treated groups compared with those in the control. There was a significant increase in apoptotic cell death in high dose treated group compared with low dose treated and control groups. No significant difference in serum testosterone level was observed after one month in the three groups. Conclusion: Buserelin induces apoptotic cell death and decreased diameter and epithelium thickness of seminiferous tubules in the adult rat testes.

Needle Aspiration Biopsy of Seminiferous Tubules (NABST) as a Diagnostic Method in Korea / 전남의대학술지

To evaluate the diagnostic efficacy of needle aspiration biopsy of seminiferous tubules (NABST) and to represent the redistributed diagnostic results corresponding to testicular volumes and follicle-stimulating hormone (FSH) levels. In this retrospective study, we investigated 65 infertile men with either azoospermia or oligoasthenoteratozoospermia. Following NABST, specimens were stained with hematoxylin and eosin and classified into five histological types. With pre-procedure FSH levels and testicular volumes, we evaluated the probabilities of detecting sperms within biopsy specimens. NABST led to the classification of normal spermatogenesis in 31 cases (47.7%), hypospermatogenesis in 23 cases (35.4%), maturation arrest in 4 cases (6.2%), and Sertoli cell only syndrome in 4 cases (6.2%). The success rate of reaching a histological diagnosis using NABST was 95.4% (62 out of 65 cases). Fourteen patients (21.5%) had a testicular volume <15 cc; of these, 8 patients (57.1%) had normal spermatogenesis, 2 patients (14.3%) had hypospermatogenesis, 2 patients (14.3%) had maturation arrest and 2 patients (14.3%) had Sertoli cell-only syndrome (SCO). Twelve patients (18.5%) had an FSH level ≥10 IU; of these, 6 (50%) had normal spermatogenesis, 2 patients (16.7%) had maturation arrest and 4 patients (33.3%) had SCO. Cases with an FSH level <10 IU were positively associated with a probability of detecting sperm using NABST (p<0.001). NABST is a reliable tool for the histological diagnosis of azoospermic and oligoasthenoteratozoospermic patients. The diagnostic success rate was high and associated with pathological accuracy. NABST is a convenient procedure with few complications.

Transcriptional regulatory networks underlying the reprogramming of spermatogonial stem cells to multipotent stem cells

Spermatogonial stem cells (SSCs) are germline stem cells located along the basement membrane of seminiferous tubules in testes. Recently, SSCs were shown to be reprogrammed into multipotent SSCs (mSSCs). However, both the key factors and biological networks underlying this reprogramming remain elusive. Here, we present transcriptional regulatory networks (TRNs) that control cellular processes related to the SSC-to-mSSC reprogramming. Previously, we established intermediate SSCs (iSSCs) undergoing the transition to mSSCs and generated gene expression profiles of SSCs, iSSCs and mSSCs. By comparing these profiles, we identified 2643 genes that were up-regulated during the reprogramming process and 15 key transcription factors (TFs) that regulate these genes. Using the TF-target relationships, we developed TRNs describing how these TFs regulate three pluripotency-related processes (cell proliferation, stem cell maintenance and epigenetic regulation) during the reprogramming. The TRNs showed that 4 of the 15 TFs (Oct4/Pou5f1, Cux1, Zfp143 and E2f4) regulated cell proliferation during the early stages of reprogramming, whereas 11 TFs (Oct4/Pou5f1, Foxm1, Cux1, Zfp143, Trp53, E2f4, Esrrb, Nfyb, Nanog, Sox2 and Klf4) regulated the three pluripotency-related processes during the late stages of reprogramming. Our TRNs provide a model for the temporally coordinated transcriptional regulation of pluripotency-related processes during the SSC-to-mSSC reprogramming, which can be further tested in detailed functional studies.