Three-dimensional imaging of the developing human fetal urogenital-genital tract: Indifferent stage to male and female differentiation.

Recent studies in our lab have utilized three imaging techniques to visualize the developing human fetal urogenital tract in three dimensions: optical projection tomography, scanning electron microscopy and lightsheet fluorescence microscopy. We have applied these technologies to examine changes in morphology and differential gene expression in developing human external genital specimens from the ambisexual stage (<9 weeks fetal age) to well-differentiated male and female organs (>13 weeks fetal age). This work outlines the history and function of each of these three imaging modalities, our methods to prepare specimens for each and the novel findings we have produced thus far. We believe the images in this paper of human fetal urogenital organs produced using lightsheet fluorescence microscopy are the first published to date.

Macroscopic whole-mounts of the developing human fetal urogenital-genital tract: Indifferent stage to male and female differentiation.

We present a detailed review of fetal development of the male and female human urogenital tract from 8 to 22 weeks gestation at the macroscopic and morphometric levels. Human fetal specimens were sexed based on macroscopic identification of fetal testes or ovaries, Wolffian or Müllerian structures and the presence of the SRY gene in the specimens at or near the indifferent stage (8-9 weeks). Specimens were photographed using a dissecting microscope with transmitted and reflected light. Morphometric measurements were taken of each urogenital organ. During this time period, development of the male and female urogenital tracts proceeded from the indifferent stage to differentiated organs. The kidneys, ureters, and bladder developed identically, irrespective of sex with the same physical dimensions and morphologic appearance. The penis, prostate and testis developed in males and the clitoris, uterus and ovary in females. Androgen-dependent growth certainly influenced size and morphology of the penile urethra and prostate, however, androgen-independent growth also accounted for substantial growth in the fetal urogenital tract including the clitoris.

Pathological changes in Fenneropenaeus indicus experimentally infected with white spot virus and virus morphogenesis.

To demonstrate pathological changes due to white spot virus infection in Fenneropenaeus indicus, a batch of hatchery bred quarantined animals was experimentally infected with the virus. Organs such as gills, foregut, mid-gut, hindgut, nerve, eye, heart, ovary and integument were examined by light and electron microscopy. Histopathological analyses revealed changes hitherto not reported in F. indicus such as lesions to the internal folding of gut resulted in syncytial mass sloughed off into lumen, thickening of hepatopancreatic connective tissue with vacuolization of tubules and necrosis of rectal pads in hindgut. Virus replication was seen in the crystalline tract region of the compound eye and eosinophilic granules infiltrated from its base. In the gill arch, dilation and disintegration of median blood vessel was observed. In the nervous tissues, encapsulation and subsequent atrophy of hypertrophied nuclei of the neurosecretory cells were found. Transmission electron microscopy showed viral replication and morphogenesis in cells of infected tissue. De novo formed vesicles covered the capsid forming a bilayered envelop opened at one end inside the virogenic stroma. Circular vesicles containing nuclear material was found fused with the envelop. Subsequent thickening of the envelop resulted in the fully formed virus. In this study, a correlation was observed between the stages of viral multiplication and the corresponding pathological changes in the cells during the WSV infection. Accordingly, gill and foregut tissues were found highly infected during the onset of clinical signs itself, and are proposed to be used as the tissues for routine disease diagnosis.

Influence of the physical properties of two-dimensional polyester substrates on the growth of normal human urothelial and urinary smooth muscle cells in vitro.

Although synthetic biomaterials have a wide range of promising applications in regenerative medicine and tissue engineering, there is limited insight into the basic materials properties that influence cellularisation events. The aim of this study was to investigate the influence of the physical properties of polyester films on the adherence and growth of normal human urothelial and urinary smooth muscle (SM) cells, as part of a programme for the development of potential biomaterials for bladder tissue engineering. Films of different thickness were produced by spin coating from solution. Cell attachment and proliferation were analysed and revealed a reproducible and significant growth advantage over the initial 7 days for both cell types on poly(lactide-co-glycolide) (PLGA) versus poly(epsilon-caprolactone) (PCL), and on thick versus thin films. In order to understand the basis of the variation in cell growth, the surface morphology, degradation behaviour and mechanical properties of the films were investigated. The pattern of cell attachment and growth was found to be unrelated to surface topography and no distinction in film degradation behaviour was found to account for differences in cell growth, except at late time points (14 days), where degradation of thin PLGA films became significant. By contrast, the flexural loss and storage moduli were found to be reduced in films composed of PLGA versus PCL, and also as film thickness increased, indicating that mechanical properties of biomaterials can influence cell growth. We conclude that elastic modulus is relevant to biology at the cellular scale and may also be influential at the tissue/organ level, and is a critical parameter to be considered during development of synthetic biomaterials for tissue engineering.

L1CAM in the Early Enteric and Urogenital System.

L1 cell adhesion molecule (L1CAM) is a transmembrane molecule belonging to the L1 protein family. It has shown to be a key player in axonal guidance in the course of neuronal development. Furthermore, L1CAM is also crucial for the establishment of the enteric and urogenital organs and is aberrantly expressed in cancer originating in these organs. Carcinogenesis and embryogenesis follow a lot of similar molecular pathways, but unfortunately, comprehensive data on L1CAM expression and localization in human developing organs are lacking so far. In the present study we, therefore, examined the spatiotemporal distribution of L1CAM in the early human fetal period (weeks 8-12 of gestation) by means of immunohistochemistry and in situ hybridization (ISH). In the epithelia of the gastrointestinal organs, L1CAM localization cannot be observed in the examined stages most likely due to their advanced polarization and differentiation. Despite these results, our ISH data indicate weak L1CAM expression, but only in few epithelial cells. The genital tracts, however, are distinctly L1CAM positive throughout the entire fetal period. We, therefore, conclude that in embryogenesis L1CAM is crucial for further differentiation of epithelia.

TRPV4 channels in the human urogenital tract play a role in cell junction formation and epithelial barrier.

AIM: The molecular interactions between transient receptor potential vanilloid subtype 4 channels (TRPV4) and cell junction formation were investigated in the human and mouse urogenital tract. MATERIALS AND METHODS: A qualitative study was performed to investigate TRPV4 channels, adherence junctions (AJs) and tight junctions (TJs) in kidney, ureter and bladder tissues from humans and wild-type and transgenic TRPV4 knockout (-/-) mice with immunohistochemistry, Western blotting, immunoprecipitation and reverse trasnscription-PCR. Cell junction formation in the wild-type and TRPV4 knockout (-/-) mouse was evaluated with immunohistochemistry and transmission electron microscope (TEM) techniques. RESULTS: TRPV4 channels are predominantly located in membranes of epithelial cells of the bladder, ureter and the collecting ducts of the kidney. There is a molecular interaction between the TRPV4 channel and the AJ. TEM evaluation showed that AJ formation is disrupted in the TRPV4 -/- mouse resulting in deficient intercellular connections and integrity of the epithelium. CONCLUSIONS: TRPV4 is believed to be a mechanoreceptor in the bladder. This study demonstrates that TRPV4 is also involved in intercellular connectivity and structural integrity of the epithelium.

Morphostructural analysis of the male reproductive system and DNA barcoding in Balclutha brevis Lindberg 1954 (Homoptera, Cicadellidae).

Balclutha brevis Lindberg 1954 is an allochthonous leafhopper infesting an invasive grass, Pennisetum setaceum, in Sicily and in mainland Europe; therefore, this species could compete with populations of native species, thus contributing to the loss of biodiversity. Considering the ecological implications of B. brevis, investigations on all its biological aspects represent, therefore, a premise for further studies in applied sciences. Based on the lacking ultrastructural data about the reproductive systems of the Auchenorrhyncha, we carried out morphostructural investigations on the male reproductive system of B. brevis. Further, a first report of DNA barcoding analysis (amplification and sequencing of Cytochrome Oxidase I gene) has also been performed to characterize B. brevis compared to other congeneric species. From a morphological point of view, the male reproductive system of B. brevis has an organization comparable to the general anatomical features of most of the Auchenorrhyncha species; however, comparing our data with those concerning the different groups of Cicadomorpha, some considerations are discussed. As for the histological and ultrastructural investigations, our results show a secretory activity of the various examined structures, mainly in the lateral ejaculatory ducts and in the accessory glands. The latter, in particular, show morphostructural differences comparing the distal tract to the proximal one; moreover, the histochemical techniques showed the possible presence of a lipid component in the peculiar cytoplasmic granules found in the gland cells. The significance of these findings in the accessory glands is discussed. Finally, the ultrastructural features found in the seminal vesicles are different from those of the lateral ejaculatory ducts and are indicative of the different roles played by these structures in the organization of the spermatozoa bundles.

Distribution of neuropeptide Y Y1 receptors in rodent peripheral tissues.

Using a sensitive immunohistochemical technique, the localization of neuropeptide Y (NPY) Y1-receptor (Y1R)-like immunoreactivity (LI) was studied in various peripheral tissues of rat. Wild-type (WT) and Y1R-knockout (KO) mice were also analyzed. Y1R-LI was found in small arteries and arterioles in many tissues, with particularly high levels in the thyroid and parathyroid glands. In the thyroid gland, Y1R-LI was seen in blood vessel walls lacking alpha-smooth muscle actin, i.e., perhaps in endothelial cells of capillaries. Larger arteries lacked detectable Y1R-LI. A distinct Y1R-immunoreactive (IR) reticulum was seen in the WT mouse spleen, but not in Y1R-KO mouse or rat. In the gastrointestinal tract, Y1R-positive neurons were observed in the myenteric plexus, and a few enteroendocrine cells were Y1R-IR. Some cells in islets of Langerhans in the pancreas were Y1R-positive, and double immunostaining showed coexistence with somatostatin in D-cells. In the urogenital tract, Y1R-LI was observed in the collecting tubule cells of the renal papillae and in some epithelial cells of the seminal vesicle. Some chromaffin cells of adrenal medulla were positive for Y1R. The problem of the specificity of the Y1R-LI is evaluated using adsorption tests as well as comparisons among rat, WT mouse, and mouse with deleted Y1R. Our findings support many earlier studies based on other methodologies, showing that Y1Rs on smooth muscle cells of blood vessels mediate NPY-induced vasoconstriction in various organs. In addition, Y1Rs in other cells in parenchymal tissues of several organs suggest nonvascular effects of NPY via the Y1R.

Autoradiographic studies of androgen-binding sites in the rat urogenital sinus and postnatal prostate.

Binding sites of [3H]testosterone and [3H]dihydrotestosterone in the rat fetal urogenital sinus and postnatal prostate and vagina grown in vitro were examined by steroid autoradiography. Distinct nuclear incorporation of both androgens appeared between 14.5 and 16.5 days of gestation in rat fetuses. Nuclear labelling in the sinus was restricted to the mesenchyme surrounding the epithelium which showed no nuclear labelling. A similar distribution of labelled cells was observed in male and female sinuses up to 18.5 days of gestation. By 20.5 days of gestation, the labelling in the ventral mesenchyme of female urogenital sinuses became less intense but persisted in the mesenchyme of the dorsal sinus wall from which the vagina is formed. In the postnatal prostate, the epithelium showed nuclear [3H]testosterone labelling at 10 days coinciding with the onset of its functional differentiation. Epithelial labelling became more intensive at 4 weeks post partum while that of the mesenchyme declined. The results suggest two phases of androgen action: formation of the prostatic buds mediated by the androgen-activated mesenchyme of the fetal urogenital sinus and the differentiation of the postnatal prostatic epithelium directly stimulated by androgens.

Evidence that inhibited prostatic epithelial bud formation in 2,3,7,8-tetrachlorodibenzo-p-dioxin-exposed C57BL/6J fetal mice is not due to interruption of androgen signaling in the urogenital sinus.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) inhibits the androgen-dependent processes by which the urogenital sinus (UGS) of fetal mice forms prostatic epithelial buds. This inhibition is mediated by aryl hydrocarbon receptors in UGS mesenchyme and causes prostate lobes to develop abnormally. Experiments were conducted to test the hypothesis that TCDD inhibits prostatic budding in C57BL/6J mice by inhibiting androgen signaling. In utero TCDD exposure sufficient to inhibit budding (5 microg/kg maternal dose on gestation day [GD] 13) had no effect on testicular testosterone content on GD 16 or 18. Nor did it inhibit the conversion of testosterone to 5alpha-dihydrotestosterone (DHT) by the UGS. Both hydroxyflutamide (OH-flutamide; a competitive androgen receptor antagonist) and TCDD inhibited prostatic epithelial budding by UGSs cultured in vitro with DHT. To determine if TCDD inhibits responsiveness to androgens, primary mesenchymal cells prepared from UGSs cultured for three days with DHT were transiently transfected with an androgen-responsive reporter plasmid (MMTV-luciferase). OH-flutamide prevented DHT from increasing luciferase activity in these cells but TCDD did not. The same results were obtained when the mesenchymal cells were isolated from UGSs cultured with both DHT and TCDD. The lack of effect of TCDD on androgen-dependent gene expression was not due to inability of transfected UGS mesenchymal cells to respond to TCDD, as shown by significant increases in luciferase activity after transfection with plasmids containing CYP1A1 and CYP1B1 promoters. Finally, while OH-flutamide prevented DHT from altering androgen receptor and 5alpha-reductase type II mRNA expression in UGS organ culture, TCDD had no such effects. Collectively, these results suggest that TCDD inhibits prostatic epithelial bud formation without impairing the androgen receptor signaling pathway.

The genus Acipenser as a model for vertebrate urogenital development: the müllerian duct.

The development of the müllerian duct was studied in a total of 85 specimens of Acipenser ruthenus and Acipenser baeri in the period from 7 days after hatching through 5 years of age. Normal histology on serial sections, transmission and scanning electron microscopy and bromodeoxyuridine immunohistochemistry were applied. In Acipenser, the primary set of opisthonephric nephrons possess short nephrostomial tubules with well-developed nephrostomes. Proliferating cells from the lateral side of the slightly protruding nephrostomial lips spread out over the ceolomic surface, replace here the flat mesothelium of lateral plate origin and establish the infundibular field, consisting of cuboidal or columnar cells. At about 28 days after hatching, the primordium of the müllerian infundibulum becomes visible in the form of a pocket-like invagination within the infundibular field. This invagination is found coexisting with and located laterally to the line of intact nephrostomes. The müllerian infundibulum, therefore, does not represent the homologon of a nephrostome itself, but must be regarded as a separate and secondary structure. The müllerian duct proper has its origin in cells from the bottom of the infundibular pocket. These cells grow as a tubule with a solid tip in the caudal direction, paralleling the wolffian duct, but without a contribution of cells from the latter. In Acipenser, a müllerian duct is present also in the adult male. In males as in females, the caudal extremity of the müllerian duct generally divides into two to three smaller terminals which end in the wolffian duct at different levels, but always cranial to the urogenital sinus. In most indifferent animals and in all males of this study, the tips of the müllerian terminals are closed and covered by a thin layer of wolffian epithelium. In adult females, the müllerian ducts end with open terminals. In both sexes, the adult müllerian duct is lined by a pseudostratified columnar epithelium, consisting of ciliated, non-ciliated and basal free cells.

Establishment of the urogenital junction in the male bovine embryo: an ultrastructural study.

The ultrastructure of the developing extratesticular rete testis, the efferent ductules and the establishment of the urogenital junction were studied in bovine embryos and fetuses of 41 through 95 days post conceptionem. The efferent ductules originate as a new set of secondary mesonephric tubules from the dorsal aspect of the nephric giant corpuscle and grow in the direction of the Wolffian duct. Cytological differentiation of the efferent ductules proceeds in a proximo--distal direction. At about 50-60 days, the simple columnar epithelium of the proximal portions of the efferent ductules already consists of the two typical cell types, i.e. reabsorptive principal cells with an endocytotic apparatus and a brush-border and ciliated cells. The lumen of the proximal portion is temporarily filled with intraductular blood vessels and perivascular tissue which may represent vestigial rudiments of glomeruli associated with the efferent ductules. At 50 to 60 days, the extratesticular rete still has a blastema--like appearance and consists of irregular cells with abundant glycogen. Extensions of the extratesticular rete come into contact with the efferent ductules and create the first end-to-side anastomoses with the latter. Somewhat later, the separating basal laminas vanish and invading rete cells intermingle with the epithelium of the efferent ductules, thus establishing the urogenital junction.

Confocal microscopy: principles and applications to the field of reproductive biology.

Confocal microscopy allows analysis of fluorescent labeled thick specimens without physical sectioning. Optical sections are generated by eliminating out-of-focus fluorescence and displayed as digitalized images. It allows 3-dimensional reconstruction (XYZ) and time-analysis (XYT), thus providing unique chance to link morphology with cell function. Since images are obtained by scanning, excess illumination of the specimen and quick decrease of the fluorescent signal are avoided. Resolution obtained with a Laser Scanning Confocal Microscopy (LSCM) is theoretically better than that of a conventional microscope. The preparation of the specimen may be based on standard techniques, such as immunocytochemistry applied to fixed cells, or on staining of living cells, following the use of different fluorescent probes at the same time (colocalization). In our laboratory, we use the LSCM system Fluoview version 2.1 (Olympus) to study reproductive biology of animals and humans. We work on stainings of oocytes and blastocysts (mouse, bovine, human), and human ovarian tissues. We study mitochondrial distribution, cortical granule migration, calcium oscillations and spindle quality to link culture conditions and oocyte quality. Staining of F-actin is used to check transzonal projections (in zona pellucida) or to detect abnormalities following experimental treatment. Blastocyst quality is analyzed in sequential optical sections for microfilament organization and counting of total cell number (staining with phalloidin (actin) and picogreen (DNA). Trophectoderm and inner cell mass distribution (differential staining), apoptotic cells (TUNEL method) and viable cells (live/dead test) are also evaluated. Confocal imaging can be helpful for rapid determination of follicle density (staining with AM Calcein) and follicle morphology (picogreen) in ovarian cortical biopsies. The current review describes the principles of confocal microscopy and illustrates its applications to the field of reproductive biology by a large collection of pictures.

The ultrastructure of adult Brugia malayi (Brug, 1927) (Nematoda: Filarioidea).

The ultrastruct of the adult subperiodic Brugia malayi (Brug, 1927) within pulmonary arteries of male jirds (Meriones unguiculatus) was studied by transmission electron microscopy. The cuticle consists of 10 sublayers (2 of which are prominently banded) and a typical outer unit membrane. Evidence is presented showing that the subcuticular region of the lateral chords comprises a functional complex of basal infoldings, multivesicular bodies, and associated mitochondria, which is probably engaged in the exchange of solutes across a permeable cuticle. Microbodies with paired, prominent cores, intracisternal A-particle viruslike bodies, nonstaining glycogen patches, and other structures are also present in the lateral chords. The platymyarian somatic musculature shares some coelomyarian characteristics, e.g., apparent neuromuscular connections and prominent glycogen deposits surrounded by mitochondria and other organelles. The alimentary tract has features typical of many nematodes. The luminal segments of the male and female reproductive tracts and their germinal products, excluding microfilariae, are described. Affinities with related species are discussed.

The genus Acipenser as a model for vertebrate urogenital development: ultrastructure of nephrostomial tubule formation and of initial gonadogenesis.

The ultrastructure of the nephrostomial tubule and the gonadal crest of Acipenser ruthenus were studied by transmission and scanning electron microscopy. Opisthonephric nephrostomial tubules begin to appear at the end of the first week after hatching and are regularly segmentally arranged and completely developed in the period between 10 and 35 days after hatching. Fully differentiated nephrostomial tubules connect the vestibular area of the Malpighian corpuscle with the coelomic cavity as short, open canals. Before reaching the latter, each nephrostomial tubule widens to a funnel-like structure and ends with a round opening, the nephrostome. The outer rim of the nephrostome protrudes slightly into the coelomic cavity forming the nephrostomial lips. The nephrostomial tubule is lined by one layer of cuboidal or low-columnar cells, equipped with an apical set of long kinocilia, which point generally in the direction of the Malpighian corpuscle. The cilia of the distal funnel region, however, point toward the coelomic cavity. The cells of the nephrostomial lips preserve a lower degree of cellular differentiation than the rest, display a blastema-like appearance and proliferate by frequent mitoses. Proliferating cells of the nephrostomial lips spread out on the coelomic surface and replace the flat mesothelium of lateral plate origin here. Furthermore, cells of the nephrostomial lips also show the tendency to grow downward, giving the epithelium of the lip region a multilayered appearance. Proliferating cells of the medial nephrostomial lips associate with accumulations of germ cells and form the primordium of the gonad (gonadal crest). Within the crest, the intercellular space between the germ cells and the surrounding epithelial supporting cells is filled with a basal lamina-like, electron-dense substance and may contain short, interlocking processes of the two cell types. The large germ cells of this early period have lobulated, electron-lucent nuclei, spherical mitochondria with inclusions and abundant narrow profiles of the smooth endoplasmatic reticulum.

Study of the effects of aging on macromolecular synthesis in various organ systems using microscopic radioautography.

In order to elucidate the effects of aging on macromolecular synthesis such as DNA, RNA, proteins, glucides and lipids in various organ systems of experimental animals and humans, systematic studies using light and electron microscopic radioautography in various organ systems including skeletal, muscular, digestive, respiratory, urinary, reproductive, endocrine, circulatory, nervous and sensory systems were studied after incorporation with macromolecular precursors. The experimental animals used were mainly ddY strain mice from embryo to postnatal days 1 and 3, weeks 1 and 2 or months 1, 2 and 6 months up to 1 and 2 years senescent stages. Animals were injected with [3H]-thymidine for DNA, [3H]-uridine for RNA, [3H]-amino acids for proteins, [3H]-glucose, [3H]-glucosamine and [35S]O4 for glucides, [3H]-glycerol for lipids and some low molecular target tracers such as hormones, inorganic substances and drugs. Results demonstrate that these precursors when incorporated into various cell types in various organs showed specific patterns of macromolecular synthesis as observed in perinatal to juvenile, mature and senescent stages. These effects of aging could answer some of the questions as to how but not why we get old.

Internal anatomy and ultrastructure of the male reproductive system of the Norway lobster Nephrops norvegicus (Decapoda: Astacidea).

The Norway lobster (Nephrops norvegicus) is economically important in Europe. However, apart from the female reproductive system, very little is known about its internal anatomy. This article focuses on studying the internal anatomy and ultrastructure of the male reproductive system. This system follows the general pattern found among decapod crustaceans, with several peculiarities. Testes are composed of lobular sperm ducts in which the spermatozoa are fully constituted. The spermatozoa present three lateral arms and a long acrosome, which gives a false appearance of flagellated spermatozoa. The two testes form a double H under the heart, and the vas deferens (VD) arise from each side at the posterior edge of the double H. The main characteristic of the VD is the presence of a sphincter in the enlarged area of the distal end of the middle VD. The MVD here shows an increase in musculature of the wall as compared to the VD, which regulates the passage of the sperm cord to the distal VD (DVD) and thence to the thelycum of the female. The wall of the spermatophore is formed in the distal part of the proximal VD, which surrounds the unique sperm cord present in the VD. Isolated spermatophores are not observed in the VD. The sperm cord is pinched off during copulation by the musculature of the DVD. Then, a portion of the sperm cord is transferred from each VD to form the isolated spermatophores. The wall of the spematophores and the spermatozoa that are observed inside the thelycum have the same morphology as those observed in the VD.

Mechanical stimuli-induced urothelial differentiation in a human tissue-engineered tubular genitourinary graft.

BACKGROUND: A challenge in urologic tissue engineering is to obtain well-differentiated urothelium to overcome the complications related to other sources of tissues used in ureteral and urethral substitution. OBJECTIVE: We investigated the effects of in vitro mechanical stimuli on functional and morphologic properties of a human tissue-engineered tubular genitourinary graft (TTGG). DESIGN, SETTING, AND PARTICIPANTS: Using the self-assembly technique, we developed a TTGG composed of human dermal fibroblasts and human urothelial cells without exogenous scaffolding. Eight substitutes were subjected to dynamic flow and hydrostatic pressure for up to 2 wk compared to static conditions (n=8). MEASUREMENTS: Stratification and cell differentiation were assessed by histology, electron microscopy, immunostaining, and uroplakin gene expression. Barrier function was determined by permeation studies with carbon 14-urea. RESULTS AND LIMITATIONS: Dynamic conditions showed well-established stratified urothelium and basement membrane formation, whereas no stratification was observed in static culture. The first signs of cell differentiation were perceived after 7 d of perfusion and were fully expressed at day 14. Superficial cells under perfusion displayed discoidal and fusiform vesicles and positive staining for uroplakin 2, cytokeratine 20, and tight junction protein ZO-1, similar to native urothelium. Mechanical stimuli induced expression of the major uroplakin transcripts, whereas expression was low or undetectable in static culture. Permeation studies showed that mechanical constraints significantly improved the barrier function compared to static conditions (p<0.01 at 14 d, p<0.05 at 7 d) and were comparable to native urothelium. CONCLUSIONS: Mechanical stimuli induced in vitro terminal urothelium differentiation in a human genitourinary substitute displaying morphologic and functional properties equivalent to a native urologic conduit.

Internal anatomy and ultrastructure of the male reproductive system of the spider crab Maja brachydactyla (Decapoda: Brachyura).

The morphology and function of the male reproductive system in the spider crab Maja brachydactyla, an important commercial species, is described using light and electron microscopy. The reproductive system follows the pattern found among brachyuran with several peculiarities. The testis, known as tubular testis, consists of a single, highly coiled seminiferous tubule divided all along by an inner epithelium into germinal, transformation, and evacuation zones, each playing a different role during spermatogenesis. The vas deferens (VD) presents diverticula increasing in number and size towards the median VD, where spermatophores are stored. The inner monostratified epithelium exocytoses the materials involved in the spermatophore wall formation (named substance I and II) and spermatophore storage in the anterior and median VD, respectively. A large accessory gland is attached to the posterior VD, and its secretions are released as granules in apocrine secretion, and stored in the lumen of the diverticula as seminal fluids. A striated musculature may contribute to the formation and movement of spermatophores and seminal fluids along the VD. The ejaculatory duct (ED) shows a multilayered musculature and a nonsecretory pseudostratified epithelium, and extrudes the reproductive products towards the gonopores. A tissue attached to the ED is identified as the androgenic gland.

An ultrastructural study of the developing urogenital tract in early human fetuses.

OBJECTIVES: This study examines the gonoducts during the ambisexual stage of human fetal development to define their ultrastructural characteristics, including the origin and antomic relationship of the mesonephric and paramesonephric ducts during gonoductal development. STUDY DESIGN: The reproductive tracts from five fetuses ranging in gestational age from 35 to 45 days were processed for ultrastructural examination. The developing mesonephric ducts, paramesonephric ducts, and their surrounding mesenchyme were studied with a Phillips 300 transmission electron microscope. RESULTS: The mesonephric ducts and paramesonephric ducts have distinctive cytoplasmic and cell surface ultrastructural characteristics, as well as different morphologic patterns of epithelial-mesenchymal interaction. Cephalad portions of mesonephric ducts and paramesonephric ducts are separated by mesenchyme, but more caudal aspects move progressively closer until they are juxtaposed but separate. CONCLUSIONS: Early mesonephric ducts and paramesonephric ducts may be distinguished because of their distinctive ultrastructural features; epithelial-mesenchymal interaction may be important in their differentiation and maintenance; both gonoducts retain their morphologic identity throughout, supporting their separate origins.