Expression of calbindin-D 28k in developing and growing chick testes

Summary Calbindin, a 28-kDa vitamin D-dependent calcium-binding protein was localized immunohistochemically in developing and growing chick testes. The protein first appeared in the germinal epithelium of developing testes of the eight-day-old embryo and remained therein throughout development. Calbindin was not present in the germinal epithelium after hatching. Calbindin was next detected in the spermatogonia and spermatocytes of one-week-old and growing chick testes. Calbindin-positive spermatogonia and spermatocytes gradually increased in number and staining intensity as the seminiferous tubules further developed. A few interstitial Leydig cells were positive for calbindin from five-week-old and older chicks. Comparison of the time-course of appearance and increase in calbindin content in spermatogonia and spermatocytes with spermatogenesis in chickens suggests that calbindin may be involved in the mitotic process in spermatogonia and spermatocytes.     

Histological and histochemical studies on the testis of camel (Camelus dromedarius) during the various seasons and ages. Part II

Histological and a few histochemical studies were done on the testes of 34 camels between 4 and 20 years of age, during three seasons: the tunica albuginea of the testes was composed of collagenous and a few elastic fibers. Isolated bundles of the smooth muscle fibers were observed in the capsule. The mediastinum testis and its radiating septula into the parenchyma also consisted of mainly collagenous and a few elastic fibers. The interstitial tissue was composed of Leydig cells and a network of reticular fibers. These cells increased in volume in the rutting period from December to March, and were distorted and reduced in volume in the nonrutting period from April to November. The seminiferous tubules were 113--250 micrometer in diameter, and were lined by many layers of spermatogenic cells and a single layer of Sertoli cells. The spermatogenesis was observed as a continuous process throughout the year, particularly in the camels between 4 and 15 years of age. Almost in all the testes studied, three types of spermatogonia, two to three layers of primary spermatocytes, rarely secondary spermatocytes and spermatids in two stages were observed. The spermatozoa were present attached to the Sertoli cells as well as free in the lumen of the tubules. The oval head of the spermatozoa was characterized by the presence of an elongated pointed, strongly PAS-positive acrosome.     

Expression of calbindin-D28k in developing and growing chick testes.

Calbindin, a 28-kDa vitamin D-dependent calcium-binding protein was localized immunohistochemically in developing and growing chick testes. The protein first appeared in the germinal epithelium of developing testes of the eight-day-old embryo and remained therein throughout development. Calbindin was not present in the germinal epithelium after hatching. Calbindin was next detected in the spermatogonia and spermatocytes of one-week-old and growing chick testes. Calbindin-positive spermatogonia and spermatocytes gradually increased in number and staining intensity as the seminiferous tubules further developed. A few interstitial Leydig cells were positive for calbindin from five-week-old and older chicks. Comparison of the time-course of appearance and increase in calbindin content in spermatogonia and spermatocytes with spermatogenesis in chickens suggests that calbindin may be involved in the mitotic process in spermatogonia and spermatocytes.     

Seasonal spermatogenic cycle and morphology of germ cells in the viviparous lizard Mabuya brachypoda (Squamata,Scincidae)

We describe seasonal variations of the histology of the seminiferous tubules and efferent ducts of the tropical, viviparous skink, Mabuya brachypoda, throughout the year. The specimens were collected monthly, in Nacajuca, Tabasco state, Mexico. The results revealed strong annual variations in testicular volume, stages of the germ cells, and diameter and height of the epithelia of seminiferous tubules and efferent ducts. Recrudescence was detected from November to December, when initial mitotic activity of spermatogonia in the seminiferous tubules were observed, coinciding with the decrease of temperature, photoperiod and rainy season. From January to February, early spermatogenesis continued and early primary and secondary spermatocytes were developing within the seminiferous epithelium. From March through April, numerous spermatids in metamorphosis were observed. Spermiogenesis was completed from May through July, which coincided with an increase in temperature, photoperiod, and rainfall. Regression occurred from August through September when testicular volume and spermatogenic activity decreased. During this time, the seminiferous epithelium decreased in thickness, and germ cell recruitment ceased, only Sertoli cells and spermatogonia were present in the epithelium. Throughout testicular regression spermatocytes and spermatids disappeared and the presence of cellular debris, and scattered spermatozoa were observed in the lumen. The regressed testes presented the total suspension of spermatogenesis. During October, the seminiferous tubules contained only spermatogonia and Sertoli cells, and the size of the lumen was reduced, giving the appearance that it was occluded. In concert with testis development, the efferent ducts were packed with spermatozoa from May through August. The epididymis was devoid of spermatozoa by September. M. brachypoda exhibited a prenuptial pattern, in which spermatogenesis preceded the mating season. The seasonal cycle variations of spermatogenesis in M. brachypoda are the result of a single extended spermiation event, which is characteristic of reptilian species. J. Morphol. © 2012 Wiley Periodicals, Inc.     

Characterization of the testicular cell types present in the rat by in vivo 31P magnetic resonance spectroscopy.

Testes of vitamin A-deficient Wistar rats before and after vitamin A replacement, of rats irradiated in utero, and of control rats were investigated by in vivo 31P magnetic resonance (MR) spectroscopy. The testicular phosphomonoester/ATP (PM/ATP) ratio ranged from 0.79 +/- 0.05 for testes that contained only interstitial tissue and Sertoli cells to 1.64 +/- 0.04 for testes in which spermatocytes were the most advanced cell types present. When new generations of spermatids entered the seminiferous epithelium, this ratio decreased. The testicular phosphodiester/ATP (PD/ATP) ratio amounted to 0.16 +/- 0.06 for testes in which Sertoli cells, spermatogonia, or spermatocytes were the most advanced cell type present. When new generations of spermatids entered the seminiferous epithelium, the PD/ATP ratio rapidly increased and finally reached a value of 0.71 +/- 0.06 for fully developed testes. Taken together, specific patterns of the PM/ATP ratio, the PD/ATP ratio, and pH were obtained that were correlated to the presence of spermatogonia, spermatocytes, round spermatids, and elongated spermatids or to the absence of spermatogenic cells. Hence, a good impression of the status of the seminiferous epithelium in the rat can be obtained by in vivo 31P MR spectroscopy.     

Heat shock protein 27 expression in the human testis showing normal and abnormal spermatogenesis

Heat shock proteins (HSPs) are molecular chaperones involved in protein folding, assembly and transport, and which play critical roles in the regulation of cell growth, survival and differentiation. We set out to test the hypothesis that HSP27 protein is expressed in the human testes and its expression varies with the state of spermatogenesis. HSP27 expression was examined in 30 human testicular biopsy specimens (normal spermatogenesis, maturation arrest and Sertoli cell only syndrome, 10 cases each) using immunofluorescent methods. The biopsies were obtained from patients undergoing investigations for infertility. The seminiferous epithelium of the human testes showing normal spermatogenesis had a cell type-specific expression of HSP27. HSP27 expression was strong in the cytoplasm of the Sertoli cells, spermatogonia, and Leydig cells. Alternatively, the expression was moderate in the spermatocytes, weak in the spermatids and absent in the spermatozoa. In testes showing maturation arrest, HSP27 expression was strong in the Sertoli cells, weak in the spermatogonia, and spermatocytes. It was absent in the spermatids and Leydig cells. In Sertoli cell only syndrome, HSP27 expression was strong in the Sertoli cells and absent in the Leydig cells. We report for the first time the expression patterns of HSP27 in the human testes and show differential expression during normal spermatogenesis, indicating a possible role in this process. The altered expression of this protein in testes showing abnormal spermatogenesis may be related to the pathogenesis of male infertility.     

Testis structure and symplastic spermatid formation during spermatogenesis of pipefishes

The pipefishes Syngnathus abaster and S. acus have paired testes of atypical organization. Each testis is a hollow tube consisting of a single germinal compartment of the tubular type. During the reproductive period, the germinal epithelium consists of small spermatocysts containing spermatogonia or primary spermatocytes. Cysts of older germ cells, such as secondary spermatocytes and spermatids were never observed. Developing symplastic spermatids were found in the lumen of the tubule together with mature sperm and large droplet-containing cells. Most of the spermatids were giant cells with four nuclei at the same developmental stage. Symplastic spermatids, which presumably form by nuclear division not followed by cytokinesis, are a stage of spermatogenesis in pipefishes.     

Effect of 17 alpha-cyanomethyl-17 beta-hydroxy-estra-4, 9-dien-3-one on reproductive organs of the male laboratory mouse

The effect of subcutaneous administration (10, 15 and 20 mg/kg body weight/day, for 21 days; and 20 mg/kg body weight/day, for 28 days) of 17 alpha-cyanomethyl-17 beta-hydroxy- estra-4, 9-dien-3-one (STS 557) on the male reproductive organs of the Parkes strain mouse was investigated. The effect of the treatment on the testis was not uniform; both regressed and normal seminiferous tubules were observed in the same section of the organ. Furthermore, the histological changes observed in the seminiferous tubules in testes of STS 557--treated mice were not different in different dosage groups. In general, in moderately affected seminiferous tubules, the germinal epithelium was thin and consisted of Sertoli cells, spermatogonia, spermatocytes and spermatids; such tubules showed presence of many vacuoles in the epithelium. In severe cases, the tubules had collapsed and were lined by mainly Sertoli cells, spermatogonia and spermatocytes. The treatment also caused marked depression in motility and concentration of spermatozoa in cauda epididymidis, weight of accessory sex glands and in the levels of sialic acid and fructose in the epididymis and seminal vesicle, respectively. By 56 days of drug withdrawal, the alterations induced in the reproductive organs returned to control levels, suggesting that STS 557 treatment induces reversible alterations in the male reproductive organs of Parkes strain mouse.     

Spermatogenic cells of the prepuberal mouse: isolation and morphological characterization

A procedure is described which permits the isolation from the prepuberal mouse testis of highly purified populations of primitive type A spermatogonia, type A spermatogonia, type B spermatogonia, preleptotene primary spermatocytes, leptotene and zygotene primary spermatocytes, pachytene primary spermatocytes and Sertoli cells. The successful isolation of these prepuberal cell types was accomplished by: (a) defining distinctive morphological characteristics of the cells, (b) determining the temporal appearance of spermatogenic cells during prepuberal development, (c) isolating purified seminiferous cords, after dissociation of the testis with collagenase, (d) separating the trypsin-dispersed seminiferous cells by sedimentation velocity at unit gravity, and (e) assessing the identity and purity of the isolated cell types by microscopy. The seminiferous epithelium from day 6 animals contains only primitive type A spermatogonia and Sertoli cells. Type A and type B spermatogonia are present by day 8. At day 10, meiotic prophase is initiated, with the germ cells reaching the early and late pachytene stages by 14 and 18, respectively. Secondary spermatocytes and haploid spermatids appear throughout this developmental period. The purity and optimum day for the recovery of specific cell types are as follows: day 6, Sertoli cells (purity>99 percent) and primitive type A spermatogonia (90 percent); day 8, type A spermatogonia (91 percent) and type B spermatogonia (76 percent); day 18, preleptotene spermatocytes (93 percent), leptotene/zygotene spermatocytes (52 percent), and pachytene spermatocytes (89 percent), leptotene/zygotene spermatocytes (52 percent), and pachytene spermatocytes (89 percent).     

Ultrastructure of the testis in Synbranchus marmoratus (Teleostei,Synbranchidae): the germinal compartment

Synbranchus marmoratus, is a protogynic diandric species in which two types of males, primary and secondary, are found. In both types, the germinal compartment in the testes is of the unrestricted lobular type, but in secondary (sex reversed females) males the lobules develop within the former ovarian lamellae. In the present study, the germinal compartment was examined in both types of males using light microscopy as well as scanning and transmission electron microscopy. Germinal compartment is limited by a basement membrane and contains Sertoli and germ cells. During maturation, processes of Sertoli cells form the borders of spermatocysts containing isogenic germ cells. Characteristically, type A and type B spermatogonia have a single nucleolus and grouped mitochondria associated with dense bodies or nuage. Type B spermatogonia, spermatocytes and spermatids are joined by cytoplasmatic bridges and are confined within spermatocysts. Secondary spermatocytes are difficult to find, indicating that this stage is of short duration. Biflagellated spermatozoa have a rounded head, no acrosome, and possess a midpiece consisting of two basal bodies, each of which produces a flagellum with a typical 9+2 microtubular composition. No associations occur between sperm and Sertoli cells. There were no differences between spermatogenesis in primary and secondary males in this protogynic, diandric fish.     

The germ cell development strategy and seasonal changes in spermatogenesis and Leydig cell morphologies of the spiny lizard Sceloporus mucronatus (Squamata: Phrynosomatidae)

The viviparous lizards of the Sceloporus genus exhibit both seasonal and continuous spermatogenesis. The viviparous lizard Sceloporus mucronatus from Tecocomulco, Hidalgo, México, exhibits seasonal spermatogenesis. This study demonstrates the relationship between changes in testis volume, spermatogenesis activity, and Leydig cells during the male reproductive cycle of S. mucronatus. A recrudescence period is evident, which starts in the winter when testicular volume is reduced and climaxes in February, when the greatest mitotic activity of spermatogonia occurs. The testicular volume and Leydig cell index increase gradually during the spring with primary spermatocytes being the most abundant cell type observed within the germinal epithelium. In the summer, the secondary spermatocytes and undifferentiated round spermatids are the most abundant germinal cells. The breeding season coincides with spermiogenesis and spermiation; testicular volume also increases significantly as does the Leydig cell index where these cells increase in both cytoplasmic and nuclear volume. During fall, testicular regression begins with a significant decrease in testicular volume and germinal epithelium height, although there are remnant spermatozoa left within the lumen of the seminiferous tubules. During this time, the Leydig cell index is also reduced, and there is a decrease in cellular and nuclear volumes within these interstitial cells. Finally, during quiescence in late fall, there is reduced testicular volume smaller than during regression, and only spermatogonia and Sertoli cells are present within the seminiferous tubules. Leydig cells exhibit a low index number, their cellular and nuclear volumes are reduced, and there is a depletion in lipid inclusion cytoplasmically.     

A quantitative analysis of germ cells and the histone variants in the testes of vitamin A-deficient rats and during subsequent repletion with vitamin A

A quantitative analysis of the different types of germ cells present in the seminiferous tubules of vitamin A-deficient-retinoate maintained rats revealed that the number of pachytene spermatocytes and spermatogonia was greatly reduced in the deficient rats. Spermatids were virtually absent in the deficient tubules which contained mostly spermatogonia and preleptotene spermatocytes along with the Sertoli cells. There was no change in the number of Sertoli cells present in the tubules of deficient rats as compared to that of normal rats. Following supplementation of retinyl acetate to vitamin A-deficient-retinoate maintained rats, there was an immediate thinning of the germinal epithelium resulting from the sloughing off of the damaged spermatocytes which were beyond repair. However, after 12 days of vitamin A supplementation fresh batch of pachytene spermatocytes started appearing while by day 16 round spermatids could be seen. Analysis of the acid soluble proteins from nuclei on different types of Polyacrylamide gel electrophoretic systems has revealed that the levels of the testis specific histone variants Hlt, TH2A and TH2B, synthesized predominantly in the pachytene spermatocytes were greatly reduced in the testes of retinoate maintained rats. Following supplementation of retinyl acetate for either 4 days or 8 days the levels of these histone variants further decreased which correlated with the decrease in the number of pachytene spermatocytes. However, by day 12 of supplementation onwards, their levels started increasing and reached near normal levels by day 24 of vitamin A-supplementation     

Dehydrodolichyl diphosphate synthase in Sertoli and spermatogenic cells of prepuberal rats

The specific activity of 2,3-dehydrodolichyl diphosphate synthase in homogenates of protease-treated seminiferous tubules, enriched spermatogenic cells, and Sertoli cells changed as a function of the age of prepuberal rats. The highest enzymatic activity occurred in each case in 23-day-old rats. Homogenates of pachytene spermatocytes, spermatids, or Sertoli cells had higher synthase activity than a whole testicular homogenate prepared by protease treatment of tubules. Enzymatic activity in pachytene spermatocytes expressed per mg of protein was about 1.7-fold higher than in spermatids, 5.3-fold higher than in spermatogonia, and about 8.3-fold higher than in spermatozoa. Therefore, the increase in spermatogenic cell synthase before day 23 can be accounted for by the appearance of the pachytene spermatocytes. Enzymatic activity decreased remarkably after the differentiation of spermatids into spermatozoa. Synthase activity in enriched Sertoli cell preparations was 1.5-2.3-fold higher than in spermatogenic cell preparations between days 15 and 30. Therefore, both spermatogenic cells and Sertoli cells contribute to changes in the enzymatic activity in seminiferous tubules during development. These changes may be important in regulating the availability of dolichyl phosphate for glycoprotein synthesis during early stages of differentiation.     

Spermatogenesis and related plasma androgen levels in Atlantic halibut (Hippoglossus hippoglossus L.)

Spermatogenesis in male Atlantic halibut (Hippoglossus hippoglossus L.) was investigated by sampling blood plasma and testicular tissue from 15-39-month-old fish. The experiment covered a period in which all fish reached puberty and completed sexual maturation at least once. The germinal compartment in Atlantic halibut testis appears to be organized in branching lobules of the unrestricted spermatogonial type, because spermatocysts with spermatogonia were found throughout the testis. Spermatogenesis was characterized histologically, and staged according to the most advanced type of germ cell present: spermatogonia (Stage I), spermatogonia and spermatocytes (Stage II), spermatogonia, spermatocytes and spermatids (Stage III), spermatogonia, spermatocytes, spermatids and spermatozoa (Stage IV), and regressing testis (Stage V). Three phases could be distinguished: first, an initial phase with low levels of circulating testosterone (T; quantified by RIA) and 11-ketotestosterone (11-KT; quantified by ELISA), spermatogonial proliferation, and subsequently the initiation of meiosis marked by the formation of spermatocytes (Stage I and II). Secondly, a phase with increasing T and 11-KT levels and with haploid germ cells including spermatozoa present in the testis (Stage III and IV). Thirdly, a phase with low T and 11-KT levels and a regressing testis with Sertoli cells displaying signs of phagocytotic activity (Stage V). Circulating levels of 11-KT were at least four-fold higher than those of T during all stages of spermatogenesis. Increasing plasma levels of T and 11-KT were associated with increasing testicular mass throughout the reproductive cycle. The absolute level of, or the relation between, testis growth and circulating androgens were not significantly different in first time spawners compared to fish that underwent their second spawning season. These results provide reference levels for Atlantic halibut spermatogenesis.     

Sulfhydryl oxidase immunoreactivity in seminiferous tubules of infertile men

Summary Sulfhydryl oxidase (SOx) immunoreactivity was investigated in the seminiferous epithelium of human biopsy material from the testes of 33 adult men with disturbed fertility. SOx immunoreactivity was expressed in normal seminiferous epithelium in type-A spermatogonia (27±4% of all spermatogonia) (n=4), in spermatocytes and round spermatids. Mature spermatozoa as well as Sertoli cells were unlabelled. within the interstitium, Leydig cells were immunopositive. In biopsies of oligozoospermic men showing hypospermatogenesis (n=24), an increase in labelled spermatogonia up to more than 90% was observed in biopsies, where seminiferous epithelia revealed only spermatogonia and Sertoli cells. Within the group of oligozoospermic patients there was a significant increase of labelled spermatogonia from 43±13% (>20 mill/ejaculate) (n=7) to 55±16% ( 20 and >20 mill/ejaculate) (n=6) to 68±8% (<5 mill/ejaculate) (n=11) and a significant (P=0.01) decrease of score count from 7.0±2.7 to 2.0±1.8. In this group the increase of labelled spermatogonia was correlated with sperm concentrations in the ajaculate (correlation coefficient: r=-0.6). In biopsies of azoospermic patients showing maturation arrest at the level of spermatocytes or spermatids (n=5) the percentage of labelled spermatogonia was within the range of 24% to 59%. Immunoreactivity in Sertoli cells was only found in single degenerating cells and in tubules showing Sertoli Cell Only Syndrome (SCO) without lumen formation. Sertoli cells within immature seminiferous cords were immunonegative, indicating that Sertoli cell SOx immunoreactivity is rather a sign of physiological alterations in degenerating cells than dependent on the stage of differentiation. Leydig cells did not show changes of immunoreactivity in any biopsy. It is concluded that SOx expression in spermatogonia may serve as a marker for spermatogenic efficiency.     

Histology of salmonid testes during maturation

The commonly applied classification systems of fish gonad maturity divide the maturation process into certain stages. However, the scales do not entirely reflect the continuity of the maturation process. Based on light microscope observations, the paper describes a comprehensive pattern of testicular transformations during maturation. The study was carried out on precocious underyearling and 1-year-old males of sea trout (Salmo trutta m. trutta L.), 1-year-old males of salmon (Salmo salar L.), and males of brown trout (Salmo trutta m. fario L.) aged from 7 months to 4 years. A total of 821 gonads collected during all seasons of the year were examined. The fish were fixed in Bouin's fluid. Histological slides of the mid-part of the gonad were made using the standard paraffin technique. The 3-6 microm sections were stained with Heidenhain haematoxylin. Histological changes of testes during maturation were similar in the three species studied. Immature and resting gonads contained type A spermatogonia in lobules only. The appearance of cystic structures containing type B spermatogonia in the lobules signalled the beginning of the sexual cycle in male gonads. Type B spermatogonia underwent synchronous mitotic divisions resulting in an increase in the total number of spermatogonia. As the spermatogenesis continued, the gonads showed a gradual increase in the number of cysts containing cells at all the spermatogenetic stages: type B spermatogonia, primary and secondary spermatocytes, spermatids, and spermatozoa. The well-formed spermatozoa were released to the lobule lumen once the Sertoli cells and spermatozoa connections broke up and the cyst disappeared. This was a continuous process observed throughout the spawning season. The spermatozoa were moved to the efferent duct. While some of the germ cells were completing spermatogenesis, the lobules contained less and less cysts with type B spermatogonia, primary and secondary spermatocytes, and spermatids; eventually all the cells completed spermatogenesis. At the end of maturation, vacuoles, up to 18.9 microm in final diameter (brown trout), appeared in the Sertoli cells. The vacuoles were visible in the lobule wall epithelium for a prolonged period of time. In most salmonid individuals examined, the reproductive cycles were observed to overlap. In some fish, the preparation for another cycle began very early, i.e., at the and of preceding spermatogenesis, which had not been observed before. Gonad maturation in some males was incomplete.     

长吻鮠精巢发育的分期及精子的发生和形成

长吻鮠精巢的发育分为精原细胞增殖期、精母细胞生长期、精母细胞成熟期、精子细胞出现期,精子完全成熟期和精子退化吸收期。精巢的后1/3不产生也不贮存精子,精子的发生和形成经过精原细胞、精母细胞、精子细胞到精子的一系列过程。精原细胞有两种类型。精子无顶体,有中心粒帽,中片长,核凹窝和线粒体发达,鞭毛具侧鳍。