Testicular Duct Research Articles

Cloning and characterization of chst11 from Procambarus clarkii involved in the host immune response of white spot syndrome virus and Aeromonas hydrophila

Carbohydrate sulfotransferases 11 (chst11) is one of the enzymes that synthesize chondroitin sulfate (CS), which has extensive immune functions in vitro and plays a critical role in mediating the infection of host by pathogenic microorganisms. However, whether it has immune functions in crayfish is still poorly understood. In our previous study of transcriptome, chst11 was differentially expressed in susceptible individuals and resistant individuals of Procambarus clarkii after white spot syndrome virus (WSSV) injection. Thus, in this study, the sequence of chst11 was obtained from P. clarkii for the first time and analyzed, and the expression pattern of chst11 was investigated. Besides, the purified recombinant protein of chst11 effect in protection in WSSV infection was explored. The full length of chst11 was 1536 bp with an 831-bp open reading frame (ORF), which encoding 276 amino acids residues with a calculated molecular mass of 33.1 kDa. The chst11 contains a Sulfotransfer_2 domain, one N-glycosylation site and three O-glycosylation sites. Phylogenetic analysis results showed that chst11 had the highest similarity to Penaeus vannamei (79.93%). The expression pattern of chst11 in different tissues indicated that chst11 was expressed highest in gut, gill and hypodermis, lowest in testicular duct, periesophageal nerve and hemocytes. The chst11 had different expression patterns in different tissues when the crayfish was challenged by WSSV, Aeromonas hydrophila and CpG ODN. Recombinant chst11 protein significantly reduced the amount of WSSV copy number in hepatopancreas at 6 h and 12 h post injection compared to the control group injected with bovine serum albumin (BSA). It was found that chst11 protein enhanced the expression of peroxinectin, proPO in hepatopancreas and midgut and the C-type lectin (ctl) in hemocytes and hepatopancreas. Intramuscularly injection of juvenile crayfish with chst11 protein decreased 60% mortality compared to the control group with BSA. This study is the first report on the antiviral function of chst11 in the immune system of crustacean.

Testicular Structure and Seminal Pathway in the Yellowtail Tetra Astyanax altiparanae (Characiforms: Characidae).

This study aimed to describe testicular and its main ducts structure in the yellowtail tetra Astyanax altiparanae, contributing to the knowledge of the region in which semen is produced, storage and released, focusing mainly on the dynamic of germinal epithelium and Sertoli cells during germ cell maturation. Ten sexually mature male A.altiparanae had their testes processed according to the routine protocols to optical microscopy. Moreover, spermatic ducts and tubular compartment of the testes of three specimens were perfused with vinyl resin for gross anatomy and scanning electron microscopy. Astyanax altiparanae testes are paired organs, separated for most of their extension, joining posteriorly in a spermatic duct formed by a squamous simple epithelium. Seminiferous compartment presents anastomosing tubular type organisation, and spermatogonia spread along its extent. Spermatogenesis is of cystic type, and there is no main testicular duct. Spermatogenesis develops in 'waves', from posterior to anterior part of the gonad. Thus, while sperm is storage posteriorly, spermatogenesis keeps maturing germ cells anteriorly, making the germinal epithelium very dynamic, holding Sertoli cells that change their function as a cystic envelope to produce secretions of the seminal fluid and store sperm. Such kind of development is thought to be responsible by the high prolificacy of this species.

Localized, Reactive F-Actin Dynamics Prevents Abnormal Somatic Cell Penetration by Mature Spermatids

Spermatogenesis occurs inside a somatic cell enclosure. Sperm release, the most important final step and a target for contraceptives, has been extensively studied in fixed tissue preparations. Here, we provide a time-lapse description of the release process in Drosophila testis exvivo. We show that the spermatid tails exit the somatic enclosure and enter the testicular duct first, followed by the spermatid heads. Prior to this, individual spermatid heads attempt toinvade the head cyst cell, and on each occasion they are repelled by a rapid and local F-actin polymerization response from the head cyst cell. The F-actin assembly involves N-WASp, D-WIP, and Arp2/3 complex and dissipates once the spermatid head retreats back into the fold. These findings revise the existing spermiation model in Drosophila and suggest that somatic cells can actively oppose mechanical cell invasion attempts using calibrated F-actin dynamics insitu.

Spermiogenesis particularities of a sperm storage species: <i>Helicolenus dactylopterus</i> (Teleostei: Scorpaenidae)

SUMMARY: A study of the spermiogenesis and spermatozoa of Helicolenus dactylopterus was conducted. Females of this species have the capacity to store sperm within their ovaries, and male gametes have a considerable cytoplasmic mass surrounding their heads to survive the long period of intraovarian sperm storage. Our observations show that early spermatids are round-shaped cells and have a spherical nucleus with diffuse chromatin. The nuclear volume decreases as a result of progressive chromatin condensation during spermiogenesis, causing the nucleus to take on a U-shape. Flagellar insertion is not central to the nucleus but consistently occurs at an oblique angle towards one side of it. The flagellum is inserted into the nuclear fossa, without subsequent nuclear rotation. In mature spermatozoa, the flagellum is adjacent to the nucleus. A comparison of the spermatozoa in the testicular lobules and those in the intraovarian storage structures suggests that the increase in volume of the cytoplasmic mass may occur in the posterior region of the testis, in the testicular duct. Spermatozoa enter the ovary in groups that reach the ovarian lumen and are surrounded by the ovarian epithelium for storage in sperm storage crypts.

Advanced Pubertal Development in Two Adolescents with New SF-1 Mutations

Purpose SF-1 is involved in gonadal and adrenal development, sex differentiation and steroidogenesis. 14 different SF-1 mutations have been described. Discrepancy between a severely undervirilised phenotype and intact testicular architecture and Wolffian duct development is noticed. Most patients have undergone early gonadectomy, so little is known about testicular function during puberty. Material and Methods We present detailed hormonal and pathological studies in two adolescents, with SF-1 mutations and advanced pubertal development. Results Case 1 is a phenotypical girl referred at the age of 13 because of severe virilisation. The karyotype was 46,XY, levels of FSH and LH were elevated and testosterone was in the normal male pubertal range. A heterozygous deletion of the SF-1 gene was identified. Gonadectomy specimen showed a left inguinal testis with Leydig cell hyperplasia, normal number of germ cells and incomplete spermatogenesis, and a right abdominal testis with scarce germ cells displaying premalignant changes. Case 2 is a 14 year old 46,XY boy, born with ambiguous genitalia. Initially, a female gender was assigned, but later, he was reassigned male after evaluation by a specialised psychologist. A heterozygous SF-1 mutation was found. At the age of 13, bilateral testicular biopsies showed normal testes with normal number and maturation of germ cells. Spontaneous pubertal development started at 13.5 years, and clinical progression through puberty is hitherto normal. Penile length is increasing, gender identity is male. Conclusions SF-1 mutations present with wide phenotypic variability. However, a marked discrepancy between severe undervirilisation of external genitalia at birth, leading to female gender assignment in most cases, and the presence of intact testicular architecture and Wolffian structures is frequent. Despite failure of external virilisation during the fetal period, pubertal testis function and spermatogenesis seem to be partially conserved, at least in some patients. As in other forms of DSD, (pre)malignant changes may be present, especially in non-scrotal testes. a

Clasping behavior and the asymmetrically latitudinal structure of the testes in the male dwarf gourami Colisa lalia

During the course of spawning, the male dwarf gourami Colisa lalia exhibits a clasping behavior where the male bends his body around the female under the floating bubble nest. In this study, in order to investigate the role of this clasping behavior, we observed the mating behavior of C. lalia in aquaria and conducted examinations of the male’s reproductive system anatomically and histologically. Forcing two males to compete for spawning with a female resulted in one male becoming dominant over the other and ultimately clasping the female. The losing male did not attempt to “sneak” during spawning, and it was possible to confirm that the clasping behavior is indispensable for sperm emission and only performed by the dominant male. Although testes of teleosts generally longitudinally elongate (i.e., parallel to the body axis), the testes of C. lalia were latitudinally elongate (i.e., vertically with respect to the body axis) and located asymmetrically between the liver and gallbladder. The latitudinal structure of the testes was also confirmed histologically, as the main testicular duct develops vertically. The asymmetric testes did not appear to influence the way in which the body bent during the clasping maneuver because the male was observed to bend its body equally to the right and left during clasping. However, the testes were situated in the vicinity of the fulcrum formed when the male bends its body, suggesting that the latitudinal testes are structurally related to its clasping behavior (i.e., a mechanism analogous to that of a nutcracker). Following physical principles (i.e., levers and mechanical advantage), it can be proposed that male dwarf gourami squeeze semen out from their latitudinal testes by bending their bodies.

Structure of the seminal pathway in the European chub,Leuciscus cephalus (Cyprinidae); Teleostei

The testicular efferent duct system of Leuciscus cephalus (Cyprinidae), is described for three phases of testicular development. Testicular main ducts were analyzed by means of conventional histology and transmission electron microscopy. Additional techniques were applied for lectin histochemistry to determine secretory activity, as well as immunohistochemistry for cell proliferation activity and for muscle actin to demonstrate the distribution and amount of contractile cells. The contribution of the main ducts' epithelia and of degenerating spermatocytes to seminal fluid composition was confirmed, with the former being a source of carbohydrates and the latter that of phospholipids. The apical glycocalyx of epithelial cells, which is important in cell recognition and potentially involved in sperm storage, was marked by RCA I, LCA, and WGA lectin. Higher numbers of proliferating epithelial cells were ascertained during spawning phase compared to pre- and postspawning phases. In the ducts' stroma, a large number of cells expressed muscle actin and tropomyosin, indicating the ducts' contractile potential for the transport of seminal fluid towards release. Adjacent to these contractile cells, numerous nerves were found, indicating neuronal control of sperm fluid flow.

The sperm roller: a modified testicular duct linked to giant sperm transport within the male reproductive tract

The male reproductive tracts of Drosophila display considerable variation in the relative size of their components, notably of the testes, but there are few structural differences between species. Here we report a remarkable coiled structure separating the testes from the seminal vesicles in the giant sperm species Drosophila bifurca. This evolutionary novelty, known as the ‘sperm roller’, seems to be an exaggeration in the size of the testicular duct as revealed by light and electron microscopic observations. It consists of a tubular monocellular epithelium lying on the basal laminae and muscle and conjunctive cells. The lumen of the roller contains crypts. The apical membrane of the epithelial cells presents numerous long microvilli protruding into the lumen. The sperm roller structure is probably involved in managing sperm during their transit through the male genital tract, because sperm are seen in bundles at the base of the testis, whereas they are singly rolled up by the time they enter the seminal vesicles. The hypercoiling of the individual spermatozoon within the roller probably occurs as the result of an osmotic process produced by features of the epithelial wall and the dramatically increased exchange surface. This is the first report of a specialized device of this type in Drosophila or, more generally, in insects.

Intersystemic arterial anastomoses in testicles.

Anastomoses between three main vessels of the testes (testicular, cremasteric, and efferent duct arteries) at the level of the inguinal and scrotal parts of the spermatic cord are extremely rare. The role of these anastomoses in the maintenance of collateral circulation is negligible. A complex anatomical formation located near epididymis-efferent duct connection, where the three vessels are anastomosed (intersystemic junction of testicular arteries), is highly prevalent. This formation is presented by two morphological variants: closed and open vascular rings (semiring).

Tissue-specific expression of inhibin/activin subunit and follistatin mRNAs in mid- to late-gestational age human fetal testis and epididymis.

Inhibin/activin subunit (alpha, beta A, and beta B) immunoreactive protein localization patterns and cell type specific inhibin alpha-subunit mRNA expression have been examined in early- to midgestational age human fetal testes. The scarcity of available third trimester human fetal tissue has, however prevented a complete examination throughout the gestational period and the cell specific expression of follistatin and beta A- and beta B-subunit mRNAs are currently unknown at any gestational age. In the present study, this gap is filled and report mRNA expression patterns of inhibin/activin subunits in mid- and late-gestational age (21-33 wk) human fetal testes and testicular duct system. We also report the first examination of follistatin mRNA signals in the human fetal gonad is also resent in both tubular and interstitial cells, and beta B-subunit mRNA is expressed in seminiferous tubules, in mid- and late-gestational age human fetal testes. Inhibin/activin beta A-subunit mRNA was detected in the interstitial cells of remarkably well preserved mid (21 and 22 wk) and late (29 wk) gestational age testes, and is the only activin-system factor mRNA also expressed in tissue of the duct system of the testis (smooth muscle cells of the epididymis). Follistatin mRNA signal was equal to background levels in testicular and duct tissues at all ages examined. These cell specific expression patterns suggest prominent and possibly differential roles for the inhibins and activins, unopposed by gonadal follistatin, in the human fetal male reproductive system.

Stimulation of epidermal growth factor gene expression during the fetal mouse reproductive tract differentiation: role of androgen and its receptor.

We have shown previously that epidermal growth factor (EGF) plays a role in testosterone-dependent fetal Wolffian duct differentiation. To further assess the role for EGF, we determined whether EGF gene expression was modulated in response to male reproductive tract differentiation. The expression of EGF messenger RNA (mRNA) was measured by an RT-PCR assay using primer pairs spanning the coding sequence of 3228 nucleotide (nt) to 3401 nt. Using the RT-PCR reaction, an amplimer of the expected size, 173 bp, was detected in the fetal male reproductive tract. The amplimer hybridized with a radioactive probe representing an internal sequence of the amplified product and was digested by the restriction enzyme HaeIII, which has an unique cleavage site at 3365 nt. The level of EGF mRNA at different stages of sexual differentiation was measured by a newly developed quantitative competitive RNA PCR (QCPCR) assay for EGF mRNA. The assay was sensitive and reproducible within a linear range of amplification with 5 x 10(4) to 140 x 10(4) copies of mRNA. Using the quantitative competitive RNA PCR we found that the level of EGF mRNA was higher in the male reproductive tract than that in the female reproductive tract. Exposure to testosterone (40 mg/kg.day) during days 13-17 of gestation induced the Wolffian duct in the female fetuses and resulted in stimulation of EGF-mRNA expression. Similarly, an antiandrogen receptor, flutamide (100 mg/kg.day) exposure during days 13-17 of gestation inhibited male reproductive tract differentiation and resulted in inhibition of EGF-mRNA expression. Moreover, during the differentiation of the male reproductive tract there was a biphasic increase in the level of EGF-mRNA, first at day 14 of gestation, the period of onset of testicular activity and Wolffian duct morphogenesis, and second at day 18 of gestation, corresponding to onset of differentiation of the urogenital sinus, epididymal duct, and seminal vesicle. Thus, it appears that testosterone-induced male sexual differentiation is accompanied by an increase in EGF gene expression.

Onset of spermatozoal degeneration in low-fertility Delaware roosters and test for autoimmune basis.

The objectives of this study were 1) to determine the onset of a heritable reproductive disorder in the rooster that is characterized by extensive spermatozoal degeneration within the ductus deferens, and 2) to determine if autoimmunity was associated with spermatozoal degeneration. Seventy-five percent of the affected roosters did not ejaculate large percentages of degenerate spermatozoa at 20 wk of age, approximately the age of sexual maturity. Rather, seminal quality gradually declined over the next 6 wk, as both ejaculate volume and number of spermatozoa ejaculated increased. The evaluation of testicular and excurrent duct tissues via immunofluorescence failed to reveal either IgY or IgA associated with spermatozoa. While histological examination revealed greater lymphocyte numbers (P less than .05) in the proximal ductus deferens, these cells were not associated with spermatozoa nor spermatozoal clumping. While spermatozoal degeneration tends to be latent at the onset of semen production, it does not appear to be due to spermatozoal autoimmunity.

The mediastinum of the bovine testis.

The bovine testis has a central mediastinum consisting of longitudinally oriented rete channels and spacious lymph vessels, embedded in the mediastinal stroma. The latter represents a contractile-elastic unit and is composed of myofibroblasts, collagen bundles and accumulations of elastin, connecting the myofibroblasts. The dimension of the mediastinum varies in cross sections at different levels between 3.5 and 31.8 mm2. In one cross section approximately 30 rete channels and approximately 30 openings of straight testicular tubules are encountered. Nearly 25% of the area is occupied by thin-walled, valveless lymph vessels. Arterial convolutes, interpolated between straight centripetal and straight centrifugal branches of the testicular artery flank the rete on all sides. It is concluded that the pulsation within these convolutes together with the contractile-elastic stroma promotes lymph and rete content in a caudo-cranial direction. Chordae retis as described by Roosen-Runge and Holstein (1978) for the human testis are a common feature in the bovine mediastinum testis. The rete channels are lined by a simple cuboidal or columnar epithelium. Short intraepithelial crypts are present and function as epithelial reserve for dilatation and expansion of the rate. The inventory of organelles is rather inconspicuous in the rete epithelium. The apical border bears short microvilli and gives a strong reaction for alkaline phosphatase. The basal cytoplasm contains many small to medium-sized electron-dense bodies and is site of a strong acid phosphatase reaction. The rete epithelium as a whole reacts strongly with leucine aminopeptidase, the marker enzyme of the testicular excurrent duct system. Many free mononuclear cells, mostly macrophages, are observed in the basal half of the rete epithelium.

Concentration of steroid hormones and androgen binding protein (ABP) in rabbit efferent duct fluid.

Androgenic hormones are necessary for the completion of normal spermatogenesis (1) and may mediate the stimulatory influence of the gonadotrophins FSH and LH on germ cell development (2). Although the stimulatory effects of LH on Leydig cell secretion are well documented, the mechanism by which FSH influences the various testicular compartments has been more speculative. The demonstration of a high affinity androgen binding protein (ABP) in the testicular efferent duct fluid (EDF) of the rat (3,4) and rabbit (5) and its control by FSH (2,6,7) has raised the possibility that FSH may influence the local concentration of androgens in the immediate vicinity of the developing and maturing germ cells. In order to test this hypothesis, a radioimmunoassay technique, which enables the simultaneous measurement of 8 steroids, including the major androgens, (8,9) was applied to a study of the steroid concentrations in rabbit EDF. It was hoped that steroid assays in conjunction with ABP measurements in the same samples would answer the following questions: (a) What is the steroid composition of the fluid bathing the testicular sperm? (b) Is there a relationship between the concentration of androgens and the levels of ABP?

THE OCCURRENCE OF HISTOCHEMICAL ACTIVITY OF 3β-HYDROXYSTEROID DEHYDROGENASE IN THE DEVELOPING TESTES OF POECILIA RETICULATA.

In the mature testes of the guppy, Poecilia reticulata, some groups of cells, distributed sparsely in the interspace between the peripheral germ cell layer and the hilar duct system, show evident histochemical response for Δ5 -3β-hydroxysteroid dehydrogenase (3β-HSD). In the testis of newly delivered guppies, somatic cells are present in the testicular hilus as a compact mass without revealing any structural differentiation. In the testis of juvenile fish of the 8mm stage about 7 days after birth, interstitial cells resembling histologically those of adult testes become differentiated from the somatic cell mass and, though only in some specimens, coincidentally begin to display weak but obvious histochemical response for 3β-HSD. Thereafter the occurrence of enzyme activity becomes increasingly regular in the developing testes, and attains the adult pattern of distribution in testes of all specimens after the 11 13mm stage or 17 ∽ 20 days of age. The appearance and enhancement of 3β-HSD activity in the testis is concurrent with the differentiation and development of the testicular duct system. Treatments of newly delivered fish with methyltestosterone (30 ∽ 50 μg/g diet) distinctly stimulate the development of the duct system, which suggests a possible role of androgen secretion occurring in the early phase of the testicular development in the control of testicular organogenesis in the guppy.

Coiling of the flagellar shaft in the head region of the mature sperm of Nucella (purpura) lampillus (L.).

ABSTRACT The sperm of Nucella are long and threadlike. The fiagellar shaft runs from the acrosome at the front of the head to the tip of the tail. Its anterior portion, the head shaft, is ensheathed by the nucleus. If a suspension of sperm in normal-concentration sea water is observed under phase contrast, and allowed to dry out slowly, the nuclear material of sperm near the edge of the coverglass swells and the head shafts of these sperm are thrown into gentle spirals within the nuclei. In some sperm the nuclear material disperses completely, and the front three-quarters of the head shaft springs into a tight right-handed coil of 5–7 turns. Instantaneous coiling of the head shaft may also be induced by treatment of the sperm with x 2 concentrated sea water or 0·01 % (w/v) solution of sodium lauryl sulphate in sea water. The enzymes pronase and trypsin at a concentration of 50 μg/ml in sea water at pH 8·0 cause dispersion of the head nucleoprotein and subsequently the head shaft forms a loose coil. The appearance and activity of the sperm do not change perceptibly over a pH range of 5·5—8·5. Sperm spread on a Langmuir trough containing normal concentration sea water were negatively stained with phosphotungstic acid and examined with an electron microscope. After such treatment the nuclei are partially spread and the fibrils of the head shafts appear twisted as the wires of an electrical flex. The twisted flagellar fibrils are seen more clearly in negatively stained head shafts of sperm whose nuclei have been completely dispersed by spreading on a Langmuir trough containing × 2 concentrated sea water. Negatively stained preparations of sperm treated with enzymes show sperm with twisted and coiled head shafts. The nucleoprotein of these sperm is disaggregated into strips or sheets, or may appear as a mass of branching fibres. The flagellar fibres of the enzyme-treated sperm are often bent or broken. Longitudinal sections of mature sperm heads show that in the intact sperm the fibres of the head shaft are not twisted but run straight throughout the length of the head. Sections from the testis and testicular duct show that microtubules are present in sperm in the testis but absent from mature sperm in the testicular duct. It is suggested that the head shaft, as a consequence of some event in spermiogenesis, has an inherent tendency to twist and coil but in the mature sperm it is ‘strait-jacketed’ by the sperm nucleus.

Spermatogenesis and the structure of the mature sperm in Nucella Lapillus (L)

ABSTRACT The testis of Nucella consists of numerous tubules, all directed inwards and joining to form a common testicular duct. In a single tubule the spermatogonia lie round the periphery. Mature sperm line the lumen of the tubule. Cells in the same stage of spermatogenesis are grouped together and all members of a group pass through spermatogenesis in phase. Staining with fast green before and after treatment with Van Slyke reagent indicates a change from lysine-rich to arginine-rich histone in the maturing spermatid. Sperm of Nucella are motile throughout their length. The sperm are thread-like and about 80 μ long. The head is Feulgen-positive and about 40 μ long. The mid-piece lies behind the head and is about 8 μ long. The flagellum runs from the front end of the head to the tip of the tail; in the head it is completely surrounded by the nucleus. The spermatogonia contain two centrioles situated near the nucleus and a conspicuous Golgi complex. There are synaptinemal complexes in spermatocyte nuclei in the synapsis stage. In the early spermatid the centriole pushes a tube through the nucleus. This tube is lined by nuclear membrane and is occupied by the anterior portion of the flagellar shaft. The nucleus elongates and the nucleoprotein condenses into strands arranged helically along the long axis of the nucleus. These strands fuse to form lamellae, which disappear in the mature sperm. Mitochondria aggregate at the base of the early spermatid nucleus and form a loose spiral around the flagellar shaft. The outer mitochondrial membranes fuse. The mid-piece of the mature sperm consists of a large tubular mitochondrion enclosing a portion of the flagellar shaft. At the early spermatid stage a pro-acrosomal granule is formed from a large Golgi complex. From this the acrosome develops; it consists of a cone and an acrosome granule. There are two sets of microtubules associated with the acrosome, one lying within the cone, the other outside the cone and separated from it by a ‘ragged membrane’. The microtubules of the outer set extend backwards along the head for two-thirds of its length. The centriole which gives rise to the flagellar shaft lies at the anterior end of the head and is separated from the acrosome by a thin layer of nucleoprotein and a double layer of nuclear envelope. There is no second centriole or derivative thereof in the mature sperm. In the tail groups of coiled fibres are associated with each pair of the peripheral flagellar fibrils.