Parapodial glandular organs in Owenia borealis (Annelida: Oweniidae) and their possible relationship with nephridia.

Oweniidae is a basal group of recent annelids and nowadays it attracts the attention of researchers of many biological fields. Surprisingly, details of their anatomy, like the adult excretory system, remain obscure. Researchers recently suggested that the paired organs of tubeworms in the family Oweniidae are related to nephridia. In the current study of Owenia borealis adults, we determined that these structures are parapodial glandular organs (PGOs) and are located in the first two segments of adults. The PGOs are complex subepidermal multicellular glands that contain secretory cells, that is, goblet cells, which are differentiated by the type of the producing tube matter. The goblet cells are surrounded by muscles that are used to extrude material stored in the PGO's lumen into the external environment. The anterior pair of PGOs have very well-developed rough endoplasmatic reticulum in the proximal cells, spacious Golgi complexes, numerous nail-shaped microvilli, and apocrine secretory processes in the goblet cells of the distal parts. The posterior pair of PGOs only consists of cells, which probably produce proteinaceous fibrils. We discuss the homology of goblet cells with specific nail-shaped microvilli that produce ß-chitin within annelids. We also discuss the possibility that PGOs and nephridia have a common origin. This study provides new information on the ultrastructure of cells that secrete the organic material used to form the tubes inhabited by tube-dwelling annelids.

Interstitial cell modulation of pyeloureteric peristalsis in the mouse renal pelvis examined using FIBSEM tomography and calcium indicators.

Typical and atypical smooth muscle cells (TSMCs and ASMCs, respectively) and interstitial cells (ICs) within the pacemaker region of the mouse renal pelvis were examined using focused ion beam scanning electron (FIB SEM) tomography, immunohistochemistry and Ca2+ imaging. Individual cells within 500-900 electron micrograph stacks were volume rendered and associations with their neighbours established. 'Ribbon-shaped', Ano1 Cl- channel immuno-reactive ICs were present in the adventitia and the sub-urothelial space adjacent to the TSMC layer. ICs in the proximal renal pelvis were immuno-reactive to antibodies for CaV3.1 and hyperpolarization-activated cation nucleotide-gated isoform 3 (HCN3) channel sub-units, while basal-epithelial cells (BECs) were intensely immuno-reactive to Kv7.5 channel antibodies. Adventitial to the TSMC layer, ASMCs formed close appositions with TSMCs and ICs. The T-type Ca2+channel blocker, Ni2+ (10-200 µM), reduced the frequency while the L-type Ca2+ channel blocker (1 µM nifedipine) reduced the amplitude of propagating Ca2+ waves and contractions in the TSMC layer. Upon complete suppression of Ca2+ entry through TSMC Ca2+ channels, ASMCs displayed high-frequency (6 min-1) Ca2+ transients, and ICs distributed into two populations of cells firing at 1 and 3 min-1, respectively. IC Ca2+ transients periodically (every 3-5 min-1) summed into bursts which doubled the frequency of ASMC Ca2+ transient firing. Synchronized IC bursting and the acceleration of ASMC firing were inhibited upon blockade of HCN channels with ZD7288 or cell-to-cell coupling with carbenoxolone. While ASMCs appear to be the primary pacemaker driving pyeloureteric peristalsis, it was concluded that sub-urothelial HCN3(+), CaV3.1(+) ICs can accelerate ASMC Ca2+ signalling.

Pathological feature and immunoprofile of cystitis glandularis accompanied with upper urinary tract obstruction.

OBJECTIVE: To explore the pathological feature and immunoprofile of immunoprofile accompanied with upper urinary tract obstruction and the immunoprofile in various types of glandular cystitis. METHODS: Pathological sections from 31 cases of cystitis glandularis with upper urinary tract obstruction and 34 cases of cystitis glandularis without upper urinary tract obstruction were observed as pathological feature on microscopy. Meanwhile, an immunohistochemical analysis was employed to determine the expression of p53, Ki67, p21, MMP-9, MUC1, MUC2, and COX-2. RESULTS: In the two groups, main pathological type was transitional epithelial, followed by intestinal epithelial; other types were a few, and the difference between the two groups was not significant. All immunohistochemical expressions of p53, Ki67, p21, MMP-9, MUC1, MUC2, and COX-2 were positive in varying degrees, and there was no significant difference between the groups. Transitional epithelial type was compared with mixed type; the difference of COX-2 was significant, P < 0.05. The differences of immunohistochemical expression among other different pathologic types were not significant. CONCLUSIONS: It is suggested that glandular cystitis accompanied with upper urinary tract obstruction shares the same pathological feature and immunoprofile as that without upper urinary tract obstruction. No significant differences of immunohistochemical expression in tissue are in cystitis glandularis with different pathological types.

Telocytes in the urinary system.

BACKGROUND: Telocytes, a new type of interstitial cells, have been identified in many organs in mammals. The present studies aimed at investigating the ultrastructure, distribution and interactions of telocytes with surrounding cells in the urinary system of rats, to confirm the existence of telocytes in kidneys, ureter and urinary bladder. METHODS: Samples of kidney, ureter, or urinary bladder were harvested for the ultrastructure by the electron microscope. The primary culture of telocytes was performed to investigate the dynamic alterations. RESULTS: Telocytes mainly located in the sub-capsular space of kidney, or between smooth muscle bundles and in the lamina propria of ureter and urinary bladder. Telocytes established numerous contacts with macrophages in the sub-capsular space of kidney, or with smooth muscle cells, nerve endings as well as blood capillaries in the ureter and urinary bladder. The complete morphology of telocytes with telopodes was observed clearly through the primary cell culture from the kidney tissues of rats. CONCLUSIONS: Our data evidenced the existence of telocytes in the urinary system, which may contribute to the tissue reparation and regeneration.

Morphology and ultrastructure of the nephridial system of Hypania invalida (Grube, 1860) (Annelida, Polychaeta, Ampharetidae).

The excretory organs of the freshwater polychaete Hypania invalida have been examined using scanning and transmission electron microscopy. Three pairs of macroscopically and ultrastructurally different nephridia are present in the thorax. Intersegmental septa in the thorax are absent, with the exception of a single diaphragm between second and third chaetiger. The first pair of nephridia is anterior to this septum, the second pair crosses the septum, with the nephrostomes anterior and the ducts and the nephridiopori posterior to it, and the third pair of nephridia is entirely posterior to the diaphragm. The first two pairs of nephridia have ciliated nephrostomes of moderate size and long nephridial ducts that extend the length of the thorax. In contrast, the third pair is characterized by short ducts and very prominent nephrostomes. Macroscopically, seven different sections of nephridial duct cells can be distinguished along the length of the first two pairs of nephridia, whereas, on an ultrastructural basis, only six different regions can be identified. Only two regions of different duct cells can be recognized in the third pair of nephridia. Cells of the two anterior pairs of nephridia show typical characteristics of transport epithelia and most likely function as excretory organs. In contrast, the duct cells of the third pair are not that much differentiated and might primarily be responsible for the release of sexual products, as sperm was observed passing through these ducts. Podocyte-like cells were observed to accompany nephridial ducts.

Tamm-Horsfall protein and urinary exosome isolation.

Urinary exosomes have been proposed as starting material for discovery of protein biomarkers of kidney disease. Current protocols for their isolation use a two-step differential centrifugation process. Due to their low density, exosomes are expected to remain in the low-speed (17,000 x g) supernatant and to sediment only when the sample is spun at high speed (200,000 x g). Analysis using western blot and electron microscopy found that urinary exosomes are also present in the low-speed pellet entrapped by polymeric Tamm-Horsfall protein, thus diminishing the procedure's reproducibility. Here we show that addition of dithiothreitol to the low-speed pellet disrupted the polymeric network, presumably by reduction of disulfide bonds linking the monomers. This modification shifted the exosomal proteins from the low- to the high-speed pellet. Also, by shifting the Tamm-Horsfall protein to the high-speed pellet, the use of dithiothreitol makes it feasible to use Tamm-Horsfall protein to normalize excretion rates of exosomal proteins in spot urines. We tested this by western blot, and found that there was a high degree of correlation between exosomal proteins and Tamm-Horsfall protein in the high-speed pellet. Since the yield of exosomes by differential centrifugation can be increased by chemical reduction, Tamm-Horsfall protein may be a suitable normalizing variable for urinary exosome studies when quantitative urine collections are not practical.

Differentiation of epithelial cells in the urinary tract.

Uroplakins, cytokeratins and the apical plasma membrane were studied in the epithelia of mouse urinary tract. In the simple epithelium covering the inner medulla of the renal pelvis, no uroplakins or cytokeratin 20 were detected and cells had microvilli on their apical surface. The epithelium covering the inner band of the outer medulla became pseudostratified, with the upper layer consisting of large cells with stalks connecting them to the basal lamina. Uroplakins and cytokeratin 20 were not expressed in these cells. However, some superficial cells appeared without connections to the basal lamina; these cells expressed uroplakins Ia, Ib, II and III and cytokeratin 20, they contained sparse small uroplakin-positive cytoplasmic vesicles and their apical surface showed both microvilli and ridges. Cytokeratin 20 was seen as dots in the cytoplasm. This epithelium therefore showed partial urothelial differentiation. The epithelium covering the outer band of the outer medulla gradually changed from a two-layered to a three-layered urothelium with typical umbrella cells that contained all four uroplakins. Cytokeratin 20 was organized into a complex network. The epithelium possessed an asymmetric unit membrane at the apical cell surface and fusiform vesicles. Umbrella cells were also observed in the ureter and urinary bladder. In males and females, the urothelium ended in the bladder neck and was continued by a non-keratinized stratified epithelium in the urethra in which no urothelial cell differentiation markers were detected. We thus show here the expression, distribution and organization of specific proteins associated with the various cell types in the urinary tract epithelium.

Covert operations of uropathogenic Escherichia coli within the urinary tract.

Entry into host cells is required for many bacterial pathogens to effectively disseminate within a host, avoid immune detection and cause disease. In recent years, many ostensibly extracellular bacteria have been shown to act as opportunistic intracellular pathogens. Among these are strains of uropathogenic Escherichia coli (UPEC), the primary causative agents of urinary tract infections (UTIs). UPEC are able to transiently invade, survive and multiply within the host cells and tissues constituting the urinary tract. Invasion of host cells by UPEC is promoted independently by distinct virulence factors, including cytotoxic necrotizing factor, Afa/Dr adhesins, and type 1 pili. Here we review the diverse mechanisms and consequences of host cell invasion by UPEC, focusing also on the impact of these processes on the persistence and recurrence of UTIs.

Structural and ultrastructural studies of the urinary tract of mice inoculated with Lactobacillus fermentum.

OBJECTIVE: To assess, using structural and ultrastructural studies of the urinary tract, the effects of the intraurethral inoculation of lactobacilli (probiotic treatment) as lactobacilli are the predominant micro-organisms of the urogenital tract of humans, monkeys and mice. MATERIALS AND METHODS: Previous work showed the protective effect of Lactobacillus fermentum CRL 1058 intraurethrally inoculated against the challenge of uropathogenic Escherichia coli. There was also an effect of oestrogens and antibiotics in the kinetics of colonization of both micro-organisms in mice. In the present study L. fermentum was inoculated with agarose beads (107 colony-forming units) and the number of micro-organisms determined by plating in selective media, giving a high degree of colonization in all the organs studied. The urinary tract organs were processed by histological and electron microscopy techniques standardized in our laboratory. RESULTS: The intraurethral inoculation of lactobacilli produced no adverse effects or significant changes in any of the organs assessed (kidney, ureter, bladder or urethra), when evaluated by histological and ultrastructural techniques. CONCLUSION: The use of lactobacilli as a probiotic treatment is probably safe.

The role of the ELAV homologue EXC-7 in the development of the Caenorhabditis elegans excretory canals.

The exc mutations of Caenorhabditis elegans alter the position and shape of the apical cytoskeleton in polarized epithelial cells. Mutants in exc-7 form small cysts throughout the tubular excretory canals that regulate organismal osmolarity. We have cloned the exc-7 gene, the closest nematode homologue to the neural RNA-binding protein ELAV. EXC-7 is expressed in the canal for a short time midway through embryogenesis. Cysts in exc-7 mutants do not develop until several hours later, beginning at the time of hatching. We find that the first larval period is when the canal completes the majority of its outgrowth, and adds new apical cytoskeleton at a rapid rate. Ultrastructural studies show that exc-7 mutant defects resemble loss of beta(H)-spectrin (encoded by sma-1) at the distal ends of the excretory canals. In addition, exc-7 mutants exhibit synergistic excretory canal defects with mutations in sma-1, and EXC-7 binds sma-1 mRNA. These data imply that EXC-7 protein may affect expression of sma-1 and other genes to effect proper development of the excretory canals.

A novel linker histone-like protein is associated with cytoplasmic filaments in Caenorhabditis elegans.

The histone H1 complement of Caenorhabditis elegans contains a single unusual protein, H1.X. Although H1.X possesses the globular domain and the canonical three-domain structure of linker histones, the amino acid composition of H1.X is distinctly different from conventional linker histones in both terminal domains. We have characterized H1.X in C. elegans by antibody labeling, green fluorescent protein fusion protein expression and RNA interference. Unlike normal linker histones, H1.X is a cytoplasmic as well as a nuclear protein and is not associated with chromosomes. H1.X is most prominently expressed in the marginal cells of the pharynx and is associated with a peculiar cytoplasmic cytoskeletal structure therein, the tonofilaments. Additionally H1.X::GFP is expressed in the cytoplasm of body and vulva muscle cells, neurons, excretory cells and in the nucleoli of embryonic blastomeres and adult gut cells. RNA interference with H1.X results in uncoordinated and egg laying defective animals, as well as in a longitudinally enlarged pharynx. These phenotypes indicate a cytoplasmic role of H1.X in muscle growth and muscle function.

Vanilloid receptor 1 expression in the rat urinary tract.

Previous findings have shown that the capsaicin sensitivity of sensory fibres is due to the expression of a specific membrane protein, the vanilloid receptor type 1 (VR1). In the present work we studied the distribution, morphology and the neurochemical content of nerve fibres expressing this receptor in the rat urinary tract. Immunolabelling was performed against the VR1 and the positive fibres were examined by light and electron microscopy. Colocalisation of VR1 and substance P or calcitonin gene-related peptide immunoreactivities, and isolectin B4 binding, was evaluated under the confocal microscope. In addition, the effect of intravesical administration of resiniferatoxin, an ultra-potent vanilloid receptor agonist, in the receptor expression in the bladder was also studied. Numerous VR1-immunoreactive fibres were found in the mucosa and muscular layer of the entire urinary tract except the kidney. In the bladder, most fibres were also substance P- or calcitonin gene-related peptide-immunoreactive but did not bind isolectin B4. Under the electron microscope VR1 immunoreactivity was confined to unmyelinated axons and varicosities containing small clear and large dense-core synaptic vesicles. They occurred beneath or among epithelial cells or closely apposed to smooth muscle cells. Intravesical resiniferatoxin decreased VR1 immunoreactivity transiently. These data indicate that primary sensory fibres expressing VR1 are extremely abundant in the rat urinary tract and that, in contrast to the skin, they belong almost exclusively to the peptide-containing sub-population of primary afferents. As capsaicin-sensitive bladder afferents are involved in nociception and reflex micturition control, the numerous free terminal nerve endings expressing VR1 in the mucosa seem more adequate to accomplish the former function. However, the close apposition between VR1-expressing fibres and smooth muscle cells suggests that they may also encode the tonus of the muscular layer.

Immunolocalization of regulatory peptides and 5-HT in bovine male urogenital apparatus.

Specimens of testis, excurrent duct including the accessory genital glands and urethra throughout its extension were investigated in adult bovines, in order to immunohistochemically localize both the peptidergic innervation and the epithelial cell types belonging to the diffuse endocrine system (DES). Immunoreactivities to GRP, met- and leu-enkephalins, CGRP, NPY, substance P, VIP, somatostatin, beta-endorphin and 5-HT antisera were tested by means of a labelled streptavidin-biotin (LSAB) method. Such regulatory substances were found in components of the peripheral nervous system (nerve fibers in the connective and muscular tissues, sub- and intrapithelial nerve terminals, nerve cells bodies and fibers in intramural ganglia), and in epithelial endocrine/paracrine cells. Bovine urogenital apparatus is supplied by many peptide-containing nerves, which contain in many localizations GRP and enkephalins, and to a lesser extent substance P, CGRP, NPY and VIP. A thin network of peptidergic nerves distributes to the musculature of the canalicular organs and accessory glands. The prostatic complex was especially rich in peptidergic innervation, and also contained somatostatin- and 5-HT-secreting endocrine cells. In addition, 5-HT-immunoreactive endocrine cells were found in the bulbourethral gland and urethral epithelium. CGRP-ir nerves were present contacting striated muscle fibers of urethra (motor end plates). The testis was devoid of any immunoreactivity. These data are compared with those obtained in a companion study carried out the same organs in two species of Equidae (Equus caballus and Equus asinus). Different patterns of immunoreactivities can be outlined in these domestic ungulates.

Ultrastructure of the protonephridial system of the oncomiracidium of Encotyllabe chironemi (Platyhelminthes, Monopisthocotylea).

In the first electron-microscopic study of the protonephridium of a monopisthocotylean oncomiracidium, flame bulbs, capillaries, and excretory bladders of larval Encotyllabe chironemi were examined. Flame bulbs are formed by a terminal and a proximal canal cell, whose cytoplasmic processes interdigitate to form a typical weir. There are many internal leptotriches (cytoplasmic outgrowths into the lumen of the flame bulb), but no (or very few) external leptotriches. A septate junction, some surface lamellae, as well as numerous vacuoles, some of which open into the canal lumen, are found in the wall of the proximal canal. Lateral flames are present in the larger capillaries. Two excretory bladders, each with a nucleus and containing many excretory concrements, are located dorso-laterally. They open dorso-laterally through narrow ducts lined by a thick wall containing electron-dense and lucent vacuoles. Phylogenetic implications of the findings are discussed.

In vitro motility investigations and electron microscopic observations on children's upper urinary tract muscle wall.

This study was performed as a part of a longer research programme on urinary tract smooth muscle layer in children. All the children whose samples were investigated underwent surgery for urinary tract malformations. Specimens were taken from different segments of upper urinary tract during surgical intervention. Specimens were investigated by either in vitro motility tests or electron microscopy or both of them. Basic patterns of tissue strips were recorded after incubation of varying duration and then tested by administering neurotransmitter agents like noradrenaline and acetylcholine-bromide. Microstructure of samples were examined electron microscopically. Investigations were performed in order to find correlation between microarchitecture and motility patterns of urinary muscle wall. Factors influencing urinary muscle motility, characteristic features of impaired musculature and its possible regeneration are discussed too. Microhistological deteriorations inhibit spontaneous smooth muscle motility but muscle contractility proved by administering noradrenaline and acetylcholine-bromide remained in some extent. Taking into consideration that smooth muscle is able to regenerate and rebuild close contacts pediatric surgeon and urologist should spare kidney parenchyma as far as it is possible.

Ultrastructural localization of blood group antigen A in normal and neoplastic urothelium.

The subcellular distribution of the blood group antigen A in the transitional epithelium of the urinary tract and its neoplastic growths was studied using transmission immuno-electronmicroscopy. Sixty-five tissue specimens from 50 blood group A1 patients were processed according to an immunogold procedure which was optimized for preservation of both antigen and ultrastructure. The reactions were stronger in the glycocalyx of the luminal surfaces and at the interdigitating cytoplasmic processes of the cells. In the intracellular compartment the reactions were associated with tubulovesicular membrane-bound structures and with the Golgi complexes. Secretory products, intra- or extra-cellular, were also positive. The greatest variability was noted in the cell surface reactions, which were positive in 88% of normal but only 41% of neoplastic urothelial specimens. An inverse correlation was found between malignant behaviour and cell surface, but not intracellular, reactions. We conclude that, in transitional cell carcinomas, there is a quantitative defect in the processing of substance A which affects predominantly the cell surface component and may involve either the transport-insertion steps, the plasma membrane-associated glycosyltransferases or internalization of blood group antigen A.

Scanning electron microscopy of the upper urinary tract in transitional cell carcinoma of the renal pelvis.

Three nephrectomy specimens with transitional cell carcinoma (TCC) of the renal pelvis were thoroughly examined by both light and scanning electron microscopy. The tumours as well as the urothelium of the upper urinary tract were studied. In all three cases, extensive areas of the urothelium, even in places remote from the tumours, were found by scanning electron microscopy (SEM) to be covered by pleomorphic microvilli. This suggests that there is a widespread failure of differentiation of the urothelium to a much greater extent than can be appreciated by conventional light microscopy.

Immunohistochemical distribution of S-100 antigen in the urinary system of man and rat.

The immunohistochemical distribution of S-100, a protein originally isolated from the brain, has been investigated at the light and electron microscopic levels in rat and man urinary systems. In both species the antigen essentially exhibited the same location, restricted, with different degrees of staining, to certain cells in the kidney, i.e. collecting tubules, thin limbs of Henle's loop and renal papillae.

[Enkephalinergic receptors at the level of the lower urinary tract]. / Les récepteurs enképhalinergiques au niveau du bas appareil urinaire.

After describing the autonomic nervous system of the lower urinary tract and its neurotransmitters, the authors discuss the recent discovery of new endomorphine group mediators--the enkephalins. There exist enkephalin immunoreactive nerve fibers in the smooth muscles of the bladder and the prostate. This neuromediator is synthetized in the spinal cord in the body of the preganglionic neuron and is carried by the axon flow towards the intramural parasympathetic ganglions of the bladder. The physiological effects of enkephalins are similar to those of morphinics and are antagonized by naloxone. Enkephalins inhibit spontaneous or provoked contractions of the bladder by inhibiting action on the parasympathetic neurons. This property of relaxing the muscles of the bladder, in both volume and pressure, may open up new vistas for uropharmacological research.