Immunolocalization of centriole-associated striated rootlets in human submandibular gland cells with and without solitary cilia

Using an antibody specific to striated rootlets, we investigated the immuolocalization of striated rootlets in cells constituting human submandibular glands. Striated rootlets were positively stained in all cell types constituting acini, intercalated ducts, striated ducts, and interlobular ducts, but their shapes were different. The mean lengths of striated rootlets were 1.46 +/- 0.49, 3.15 +/- 1.35 and 3.99 +/- 1.02 microm in acinar secretory cells, myoepithelial cells, and columnar cells of the striated duct, respectively. The rootlets were the longest in columnar cells of the striated duct, in which paired centrioles were located in the apical cytoplasm away from nuclei. These findings suggest that striated rootlets play important roles in the positioning of centrioles in the cell. 2-8% of striated rootlets in myoepithelial cells were associated with solitary cilia, but they were not associated with solitary cilia in acinar cells and columnar cells of the striated duct. These observations suggest that striated rootlets may be associated with centrioles under normal physiological conditions, without formation of solitary cilia.     

Identification of a 195 kDa protein in the striated rootlet: its expression in ciliated and ciliogenic cells

Little is known about the molecular composition of the ciliary rootlet. We raised monoclonal antibodies to a crude preparation of striated rootlets isolated from the human oviduct, and obtained a clone (R4109) that specifically labeled the ciliary rootlets. Rootlets associated with the solitary cilium in secretory cells and fibroblasts were also labeled. R4109 identified a 195-kDa protein by immunoblotting. Ciliogenic cells in the oviduct epithelium of young mice were labeled in the globular and/or granular pattern by R4109 by immunofluorescence microscopy. Immunoelectron microscopy showed that they corresponded to fibrogranular complex and dense granule, respectively. The result demonstrated that the 195-kDa protein is a component common to the striated rootlet and dense granule, and thus suggested that dense granules are involved in the rootlet formation.     

Ultrastructure of multiciliated collar cells in the pilidium larva of Lineus bilineatus (Nemertini)

Summary The ultrastructure of the apical plate of the free-swimming pilidium larva of Lineus bilineatus (Renier 1804) is described with particular reference to the multiciliated collar cells. In the multiciliary collar cells there are several, up to 12, cilia surrounded by a collar of about 20 microvilli extending from the cells' apical surface. The cilia have the typical 9+2 axoneme arrangement and are equipped with striated caudal rootlets extending from the basal bodies. No accessary centriole or rostral rootlet were observed. Microvilli surrounding the cilia are joined in a cylindrical manner by a mucus-like substance to form a collar. In comparison with many sensory receptor cells built on a collar cell plan the multiciliary collar cells of the pilidium larva apical plate are rather simple and unspecialized. In other pilidium larvae monociliated collar cells are found in the apical plate. The possible function and phylogenetic implications of multiciliated collar cells in Nemertini are briefly discussed.List of Abbreviations a axoneme - b basal body - c cilia or flagella - d desmosome - G Golgi apparatus - m mitochondria - mf microfilaments - mu mucus - mv microvilli - n nucleus - nt neurotubules - pm plasma membrane - r rootlet - ri ribosomes - v secretory vesicles     

Depolymerization of microtubules by colcemid induces the formation of elongated and centriole-nonassociated striated rootlets in PtK(2) cells

Striated rootlets are cross-banded structures associated with the basal body, which extends the cilium. To determine whether microtubule dynamics influence the shape and distribution of striated rootlets, we have depolymerized the microtubules by colcemid and observed the rootlets by the immunohistochemical technique with the R4109 antibody that specifically reacts with a 195-kDa protein in the rootlets in PtK(2) cells. In control interphase cells, striated rootlets were observed in various profiles such as fibrillar, branched, or looped shapes and were associated with a pair of centrioles. Treatment with colcemid (0.1 micro g/ml or more) resulted in the elongation and/or structural complication of the centriole-associated rootlets and the organization of intracytoplasmic free rootlets. These changes appeared 6 h after colcemid treatment and became more prominent with time. The changes were reversible and almost disappeared 2 h after removal of the drug. Immunoelectron microscopy confirmed that the R4109 antibody decorated both centriole-associated rootlets and free rootlets. These findings indicate functional relationships between cytoplasmic microtubules and striated rootlets and the existence of rootlet-nucleating factors in the cytoplasm, in addition to centrioles.     

The ultrastructure of primary cilia in quiescent 3T3 cells

Electron microscopy was used to investigate primary cilia in quiescent 3T3 cells. As in the case of primary cilia of other cell types, their basal centriole was found to be a focal point of numerous cytoplasmic microtubules which terminate at the basal feet. There are also intermediate filaments which appear to converge at the basal centriole. Cross-striated fibers of microtubular diameter, reminiscent of striated rootlets of ordinary cilia, appear associated with the proximal end of the basal centriole. Usually less than nine cross-banded basal feet surround the basal centriole in a well-defined plane perpendicular to the centriolar axis. The ciliary shaft was found to be entirely enclosed in the cytoplasm of fully flattened cells. In rounded cells, it could be found extending to the outside of the cell. Periodic striations along the entire shaft were observed after preparing the cells in a special way. The tip of the shaft showed an electron-dense specialization. Several unusual forms of primary cilia were observed which were reminiscent of olfactory flagella or retinal rods.Using tubulin antibody for indirect immunofluorescence, a fluorescent rod is visible in the cells [18] which we demonstrate is identical with the primary cilium.     

Ultrastructure of human labial salivary glands. 3. Myoepithelium and ducts

Myoepithelial cells were present between the basal lamina and the acinar secretory cells of human labial salivary glands. In form and disposition, they resembled myoepithelial cells in the major salivary glands. Many of these cells possessed single cilia on their upper surfaces. Such cilia occasionally extended into invaginations of the overlying secretory cell. The intercalated ducts were variable in occurrence. Their epithelium ranged from columnar to squamous, and showed few signs of secretory activity. Few intralobular ducts possessed basal striations. While mitochondria were abundant in non-striated cells, they were randomly disposed in both basal and apical cytoplasm, and the basal plasmalemma showed only occasional infoldings. The paucity of true striated ducts in labial salivary glands may be responsible for the high concentration of sodium and chloride in unstimulated labial gland salivary secretions.     

Observations on the solitary cilium of rabbit oviductal epithelium: its motility and ultrastructure

Solitary cilia have been observed on rabbit oviductal epithelial cells. In tissue cultures of fimbrial epithelium of 3- and 4-day-old animals observed by phase microscopy, most of these single cilia exhibited a vortical or funnel-type movement while others had the usual to-and-fro motility. Primary cilia are usually considered immotile. Transmission electron microscopy of specifically identified single cilia revealed differences between the ciliary shafts and basal bodies of the single cilia as compared to those of mature oviductal ciliated cells. The basal body of the solitary cilium often had at least two triangular, striated, basal foot processes, lacked electron-dense satellite material around its basal end, and occasionally had striated rootlets. In contrast, the cilia of mature ciliated cells had only one basal foot, exhibited much electron-dense satellite material, and lacked rootlets. Cross sections of the single cilia showed patterns of microtubules different from the usual 9 + 2 axonemal complexes of normal cilia and included 9 + 0, 10 + 2 singlets, 7 + 2 doublets, and 8 + 1 doublet and 2 singlets; one did have the usual 9 + 2 arrangement. We postulate that the presence of more than one basal foot process may be responsible for the vortical motility observed. The primary cilia are shorter than normal cilia; the longest one measured was 1.86 micron in length, 0.28 micron in width at its base, and 0.14 micron at its tip. Based on the light-microscopic, scanning-electron-microscopic and transmission-electron-microscopic observations, such solitary cilia were observed more frequently in the oviductal tissues of the 3- to 4-day postnatal rabbits grown in tissue culture and in ovariectomized and ovariectomized/progesterone-treated adult animals than in estrous, ovulatory, or ovariectomized/estradiol-treated rabbits.     

Fine structure of probable mechano- and chemoreceptors in the caudal epidermis of the lugworm Arenicola marina (Annelida,Polychaeta)

Summary The caudal part of the lugworm Arenicola marina shows numerous epidermal papillae formed by a thick glandular epidermis in which ciliated sensory buds have been found. These buds comprise supporting cells and two types of receptors, R1 and R2, which are primary sensory cells whose axons are connected to the basiepidermal nerve plexus. The receptors possess several typical cilia projecting into the surrounding seawater, stout intracuticular microvilli filled with filaments, and they contain dense vesicles. The R1 cells, more numerous, show features of chemosensory cells. The rarer R2 cells have large striated rootlets surrounded by a dense sheath of fibrillar material and are probably mechanoreceptors. The physiological functions of these receptors are discussed.     

Golgi-cilium complex in rabbit ciliary process cells

We report here on a structural association of single cilia, via their striated rootlets, with the Golgi complex in epithelial cells and stromal fibroblasts of rabbit ciliary processes of the eye. The structure is designated a Golgi-cilium complex and its likely role in aqueous humor production is discussed.     

Epidermis and protonephridia of a free-living platyhelminth, Mesocastrada führmanni Voltz, 1898 (Rhabdocoela, Typhloplanoida): An ultrastructural study

The ultrastructure of the epidermis and the protonephridia of the free-living rhabdocoel Mesoscastrada führmanni is described. The epidermis consists of polarized cells, the nucleus located in the basal part of the cell and the mitochondria in the apical part. The surface is entirely covered by cilia anchored in the cytoplasm by horizontal and vertical striated rootlets. Cilia of the flame bulbs also have horizontal and vertical striated rootlets. The weir apparatus of the cyrtocyte is composed of a single row of ribs connected by a thin “membrane” of extracellular material. Bundles of microtubules, located in the ribs originate in the centrioles. Epidermal cells and flame bulbs of M. führmanni closely resemble those of the other Typhloplanoida examined so far.     

Morphofunctional characteristics of the striated centrosome rootlets of cultured embryonic pig kidney cells

The analysis of the centrosome structure in PK-cells has shown that the striated rootlets occur in 25-30% of the cells. Most frequently, the centrosome contains 1-2 rootlets, oriented either to the active or to the nonactive centriole. Occasionally, they occur in both the centrioles simultaneously. The presence of striated rootlets does not correlate with the presence of a primary cilium in the centrosome: there occur centrosomes with the primary cilium but without striated rootlets, and conversely: with striated rootlets but without the primary cilium.     

Morphology and ultrastructure of the venom glands of the northern pacific rattlesnake Crotalus viridis oreganus

The venom gland of Crotalus viridis oreganus is composed of two discrete secretory regions: a small anterior portion, the accessory gland, and a much larger main gland. These two glands are joined by a short primary duct consisting of simple columnar secretory cells and basal horizontal cells. The main gland has at least four morphologically distinct cell types: secretory cells, the dominant cell of the gland, mitochondria-rich cells, horizontal cells, and “dark” cells. Scanning electron microscopy shows that the mitochondria-rich cells are recessed into pits of varying depth; these cells do not secrete. Horizontal cells may serve as secretory stem cells, and “dark” cells may be myoepithelial cells. The accessory gland contains at least six distinct cell types: mucosecretory cells with large mucous granules, mitochondria-rich cells with apical vesicles, mitochondria-rich cells with electron-dense secretory granules, mitochondria-rich cells with numerous cilia, horizontal cells, and “dark” cells. Mitochondria-rich cells with apical vesicles or cilia cover much of the apical surface of mucosecretory cells and these three cell types are found in the anterior distal tubules of the accessory gland. The posterior regions of the accessory gland lack mucosecretory cells and do not appear to secrete. Ciliated cells have not been noted previously in snake venom glands. Release of secretory products (venom) into the lumen of the main gland is by exocytosis of granules and by release of intact membrane-bound vesicles. Following venom extraction, main gland secretory and mitochondria-rich cells increase in height, and protein synthesis (as suggested by rough endoplasmic reticulum proliferation) increases dramatically. No new cell types or alterations in morphology were noted among glands taken from either adult or juvenile snakes, even though the venom of each is quite distinct. In general, the glands of C. v. oreganus share structural similarities with those of crotalids and viperids previously described.     

Ultrastructure of a cephalic sensory organ in larvae of the gastropod Phestilla sibogae (Aeolidacea, Nudibrachia)

The cephalic sensory organ is a superficial sensory receptor located between the velar lobes at the level of the shell aperture. Three cell types make up this sensory area: (1) six flask-shaped cells bearing numerous cilia; (2) adjacent supporting or accessory cells which have numerous, often branched, microvilli; and (3) vacuolated cells which occupy the center of the area. The flask-shaped cells appear to be the sensory units. These cells have a deep invaginated lumen, with cilia arising from the cell surface in the lumen oriented either toward the base of the lumen or toward the epidermal surface. These cilia, some of which extend slightly above the body surface, are presumed to be non-motile, as they lack (dynein?) arms on the axonemal A tubules and lack striated rootlets. The six flask cells are in intimate contact with the underlying cerebral ganglia and axons from each cell pass into ganglionic tissue. The supporting cells may be sensory, but no direct connection with the nervous system was seen. The function of the central vacuolated cells is not known. This cephalic organ may be a derivative of the original apical tuft of the trochophore stage.     

Fine structure of the mandibular gland in Japanese field vole (Microtus montebelli)

The mandibular glands of the Japanese field vole were examined by light microscopy, and transmission and scanning electron microscopies. The acinar cells contained light and coarse secretory granules, and reacted with PAS and stained slightly with AB; they were considered to be seromucous in nature. The acinar epithelium was composed of light and dark cells containing many secretory granules. The intercalated duct cells consisted of light cells possessing a few dense granules. A few cytoplasmic crystalloides of moderate density were observed in occasional light cells. The striated ducts were comprized of two distinct portions, a secretory portion and a typical striated portion without secretory granules. The epithelium secretory portion consisted of light and dark cells containing acidophilic granules and exhibited a sexual dimorphism in these granules: The male epithelia contained the granules of low to high densities, while the female epithelia had only dense granules being smaller than those in the males. The epithelium of typical striated portion was composed of light and dark cells containing fine vacuoles and vesicles.     

Giant secretory granules in the ducts of the parotid and submandibular glands of the slow loris

The parotid and submandibular glands of a slow loris, a rare Southeast Asian primate, were obtained after the head had been perfused by fixative for a study of the brain. These tissues were processed by conventional means for electron microscopy. Glands also were obtained at autopsy from 2 other lorises, fixed by immersion in formalin, and subjected to a battery of tests for glycoconjugates. In the parotid gland, a short segment of the proximal striated duct lacks both basal striations and any sign of secretory activity. The major portion of the striated duct consists of tall cells that contain a spectrum of secretory granules, some larger than the nuclei (many granules are > 9 mum in diameter). These granules, which are delimited by a single membrane, are capable of chain exocytosis. Many of the giant granules have bundles of cytofilaments (4.5-6.5 nm) in apparent association with their surface. Occasional cells contain numerous small granules. Duct cells with or without granules lack basal striations. The granules contain neutral glycoconjugates but no acidic glycoconjugates. Some, but not all, interlobular excretory ducts also have secretory granules that run the gamut from tiny to giant. Exactly the same situation occurs in the submandibular gland. Unlike other primates, which may have duct cells that contain only a few tiny granules in their apices, the cells in both the striated and excretory ducts in the slow loris appear to be specialized for secretion rather than for transport. The biofunction of the giant granules is unknown.     

In vivo ciliogenesis in human fetal tracheal epithelium

Development of ciliated cells (CC) in the fetal human trachea was studied by light and electron microscopy in specimens obtained from 45 embryos or fetuses aged from 9 to 27 weeks of gestation (menstrual age). Four stages could be recognized during tracheal development. Up to 11 weeks (stage I), the trachea was covered with a columnar undifferentiated epithelium with abundant glycogen, apical microvilli, and primary cilia. From 12 to 18-19 weeks (stage II), centriologenesis and secondary ciliogenesis were very active, and the percentage of CC and secretory cells (SC) progressively increased. From 20 to 22-23 weeks, the density of CC was higher but, in parallel, the percentage of SC decreased (stage III). Throughout this period, the different steps of ciliogenesis could be identified in the same field, and the ciliated borders consisted of ciliary shafts with a disorderly arrangement. Megacilia were identified. Some of the preciliated cells had both cilia and secretory granules in their apical cytoplasm. After 24 weeks (stage IV), the ciliated border was apparently mature, the rootlets lengthened, and the cilia were correctly orientated. Whatever the fetal age, the density of CC was significantly higher (P less than .01) in the dorsal trachea compared to the ventral trachea. There are many similarities between animal and human ciliogenesis, but in human fetuses, most of the ciliary differentiation occurs early, during the first half of gestation. As demonstrated in experimental models, SC likely play a major role in genesis of CC during the fetal development of the human trachea.     

Fine structure of the excretory system of the deep posterior (Ebner's) salivary glands of the human tongue

Human deep posterior lingual glands (von Ebner's glands) are located beneath the circumvallate papillae. They are formed by tubuloalveolar adenomeres, intercalated ducts and excretory ducts coming together in the main excretory duct. The tubuloalveolar cells, pyramid-shaped, show large and dense secretory granules (clear cored) throughout the cytoplasm, rare basal folds and packed cisternae of rough endoplasmic reticulum (RER) at the basal pole. The columnar cells of the intercalated ducts are arranged in a monolayer. They are characterized by dense, clear-core secretory granules (mostly in the apical cytoplasm), a basal nucleus, well-developed RER and Golgi apparatus, and thin filaments distributed in supra- and perinuclear cytoplasm. Striated ducts are absent. Excretory ducts, coming together in the main duct, are lined by a bistratified epithelium. The inner layer consists of columnar cells showing bundles of tonofilaments with scarce secretory activity. The outer layer is composed of basal cells lying on the basal lamina. The main excretory duct, which opens at the bottom of the vallum, shows a stratified epithelium. The outer side is composed of 2-3 layers of malpighian cells lying on the basal lamina. The inner side consists of a single layer of cuboidal-columnar cells with dense apical granules and well-developed organelles synthesizing and condensing secretions. These cells interpolate with goblet cells, rare mitochondria-rich cells, ciliated cells and numerous small globous cells showing a clear matrix and lacking secretory granules. The cilia show a 9 + 2 microtubular structure with basal bodies provided with striated rootlets. Myoepithelial cells surround with their processes the basal portions of the secretory cells and the intercalated ducts. The conclusions concern some comparative aspects and some hypothesis on the functional role of goblet cells, ciliated cells and epithelial cells lining the different ducts, also in relation to the final secretory product.     

Vertebrate primary cilia: a sensory part of centrosomal complex in tissue cells,but a “sleeping beauty” in cultured cells?

Primary cilium development along with other components of the centrosome in mammalian cells was analysed ultrastructurally and by immunofluorescent staining with anti-acetylated tubulin antibodies. We categorized two types of primary cilia, nascent cilia that are about 1microm long located inside the cytoplasm, and true primary cilia that are several microm long and protrude from the plasma membrane. The primary cilium is invariably associated with the older centriole of each diplosome, having appendages at the distal end and pericentriolar satellites with cytoplasmic microtubules emanating from them. Only one cilium per cell is formed normally through G(0), S and G(2)phases. However, in some mouse embryo fibroblasts with two mature centrioles, bicilates were seen. Primary cilia were not observed in cultured cells where the mature centriole had no satellites and appendages (Chinese hamster kidney cells, line 237, some clones of l-fibroblasts). In contrast to primary cilia, striated rootlets were found around active and non-active centrioles with the same frequency. In proliferating cultured cells, a primary cilium can be formed several hours after mitosis, in fibroblasts 2-4 h after cell division and in PK cells only during the S-phase. In interphase cells, formation of the primary cilium can be stimulated by the action of metabolic inhibitors and by reversed depolymerization of cytoplasmic microtubules with cold or colcemid treatments. In mouse renal epithelial cells in situ, the centrosome was located near the cell surface and mature centrioles in 80% of the cells had primary cilium protruding into the duct lumen. After cells were explanted and subcultured, the centrosome comes closer to the nucleus and the primary cilium was depolymerized or reduced. Later primary cilia appeared in cells that form islets on the coverslip. However, the centrosome in cultured ciliated cells was always located near the cell nucleus and primary cilium never formed a characteristic distal bulb. A sequence of the developmental stages of the primary cilium is proposed and discussed. We also conclude that functioning primary cilium does not necessarily operate in culture cells, which might explain some of the contradictory data on cell ciliation in vitro reported in the literature.     

THE FINE STRUCTURE OF EPENDYMA IN THE BRAIN OF THE RAT

The ciliated ependyma of the rat brain consists of a sheet of epithelial cells, the luminal surface of which is reflected over ciliary shafts and numerous evaginations of irregular dimensions. The relatively straight lateral portions of the plasmalemma of contiguous cells are fused at discrete sites to form five-layered junctions or zonulae occludentes which obliterate the intercellular space. These fusions occur usually at some distance below the free surface either independently or in continuity with a second intercellular junction, the zonula adhaerens. The luminal junction is usually formed by a zonula adhaerens or, occasionally, by a zonula occludens. The finely granular and filamentous cytoplasm contains supranuclear dense bodies, some of which are probably lysosomes and dense whorls of perinuclear filaments which send fascicles toward the lateral plasmalemma. The apical regions of the cytoplasm contain the basal body complexes of neighboring cilia. These complexes include a striated basal foot and short, non-striated rootlets emanating from the wall of each basal body. The rootlets end in a zone of granules about the proximal region of the basal body, adjacent to which may lie a striated mass of variable shape. All components of the basal body complex of adjacent cilia are independent of each other.     

The ultrastructure of the sensory cells in the chemoreceptor of the ommatophore of Helix pomatia L.

Most of the sensory cells found in the chemoreceptor of the ommatophore of Helix pomatia are typical bipolar cells. The chemoreceptor is deveded by a furrow into two parts; within the ventral subdivision the layer of sensory cell bodiesis thicker than in the dorsal part. According to the differentiations of the apical surface of the dendrites, it is possible to distinguish six different classes: a) dendrites with one cilium and 75 nm thick cytofila (sometimes dendrites of identical appearance posses more than one cilium); b)dendrites with several cilial and 150 nm thick cytofila; c) dendrites with several cilia, 50 nm thick cytofila, and long, striated rootlets; d) dendrites with several cilia bur without cytofila; e) dendrites with 130 nm thick cytofila but without cilia; and f) dendrites with 65 nm thick cytofila but without cilia; dendrites of this class are the only ones with a cytoplasm more electron dense than that of the surrounding supporting cells. All these dendrites are connected to the surrounding supporting cells by terminal bars, each consisting of zonula adhaerens, aonula intermedia and zonula septata. The perikarya of the sensory cells measure approximately 15 mum by 8 mum and enclose 10 mum by 6 mum large nuclei. Axons, originating from these perikarya, extend to the branches of the digital ganglion. In the distal part of this gangloin the axons come into synaptic contact with interneurons, but in our electron micrography it was not possible to coordinate processes and synapses with the corresponding neurons.