Electron-lucent Vacuoles Research Articles

Ultrastructure of the frontal gland in Prorhinotermes simplex (Isoptera: Rhinotermitidae) and quantity of the defensive substance

The frontal gland as a sac-like organ in Prorhinotermes simplex is present only in presoldiers, soldiers, and imagoes, but exists also in nymph-soldier intercastes. The secretory epithelium consists of a single type of secretory cells adhering directly to the cuticular intima. Secretory vacuoles originate in electron dense vesicles, which are transformed into large electron lucent vacuoles. Intermediate vacuoles frequently contain lipid droplets. The frontal gland cells in presoldiers reveal modifications connected with the production of a new cuticle; the new cuticle is thin and compact, whereas the old one is thick, porous, and wrinkled. None of these cuticles is present in soldiers (sic!). In soldiers, the cuticular intima is of endocuticular origin and is formed by dispersed dense material; the apical parts of secretory cells are formed by numerous irregular finger-like projections, true microvilli are completely lacking. In imagoes, the cuticle is composed of an epicuticle, a layer of epicuticular filaments, and one more basal layer; sexual differences were not observed. In nymph-soldier intercastes, the structure of the gland differs in the head and in the metathorax; the head part of the gland resembles the imaginal gland whereas the thoracic part resembles more that of the soldier; the development of secretory vacuoles stops at the stage of presence of lipid droplets. E-1-nitropentadecene was found in the highest amount in soldiers (comparable to P. inopinatus soldiers), in moderate amount in imagoes, and it is missing in both presoldiers and intercastes.

The gastric shield and the underlying epithelium ofChlamys farreri: Morphological, histological and histochemical characterizations

The gastric shield and underlying stomach epithelium ofChlymas farreri were examined at the light and ultrastructural levels. The results showed that the gastric shield consisted of two different size lobes joined together by a narrow middle piece, the thicker lobe was shaped like a funnel, but unclosed at the lateral side; the other lobe was irregularly triangular-shaped. The transverse section of the thicker lobe was obviously laminated and gradually decreased in thickness from the peak to the margins of the shield. The underlying epithelium bore numerous about 3μm diameter spherical processes formed by the apical plasmalemma of the epithelial cells becoming blunt pseudopodia. Microvilli and some interspersed cilia were present in the areas among the spherical processes regions where only microvilli existed. Rough endoplasmic reticulum, Golgi apparatus, different-sized electron-dense secretory granules and electron-lucent vacuoles as well as abundant mitochondria were present in the underlying epithelial cells. Fused droplets of the secretion from the underlying epithelial cells formed the gastric shield.

Changes in intestinal villi, cell area and intracellular autophagic vacuoles related to intestinal function in chickens

1. Morphological changes in the intestinal villi, cell area and cell mitosis number in the duodenal epithelial cells were compared in cockerels fasted for 1, 2 and 3 d, and also when refed for 1 and 2 d after 3 d of fasting, to demonstrate whether these morphological changes are related to intestinal function. Alterations in the fine structure of vacuoles in epithelial cells were also examined in each group to investigate whether the vacuolar changes are associated with these morphological changes, and to obtain an index for judging the nutritional condition of the chicken intestine. 2. Fasting induced decreases in villus height, cell area and cell mitosis number, which recovered rapidly after refeeding, suggesting that these parameters are related to changes in intestinal function and may be useful for assessing intestinal function. 3. At 1 d of fasting, small electron-dense bodies appeared in the absorptive epithelial cells, some of them fusing with each other. As the fasting period increased, these small bodies developed to moderate-sized nascent autophagic vacuoles containing various kinds of electron-dense contents and finally became large autophagic vacuoles with electron-lucent contents. Some vacuoles showed positive acid phosphatase reactions, which indicated that they were lysosomal autophagic vacuoles containing hydrolytic enzymes. 4. After 1 d of refeeding the large autophagic vacuoles seen after 3 d fasting rapidly decreased to the small electron-dense bodies seen after 1 d of fasting. 5. These findings suggest that intestinal epithelial cells have the ability to digest their own cell components to supply nutrients during fasting by means of lysosomal active autophagic transport mechanisms; after refeeding, the epithelial cells return to the absorption of nutrients. 6.The present results demonstrate that the autophagic vacuolar changes are correlated with changes in intestinal villus height, cell area and cell mitosis number induced by fasting and refeeding; this indicates that autophagic vacuoles are a useful index of the nutritional condition of chicken intestine. The greater the number of electron-lucent vacuoles there are in the duodenal absorptive cells, the lower the nutritional condition of the chicken intestine.

Pathology of intrinsic cardiac neurons from ischemic human hearts.

Various populations of intrinsic cardiac neurons influence regional cardiac function tonically. It is not known whether such neurons are affected by disease states and, if so, in what manner. Therefore, the morphology of intrinsic cardiac ganglia obtained from patients with angiographic evidence of compromised regional coronary blood supply was studied. Posterior atrial ganglia and surrounding fat, removed at the time of cardiac surgery, were placed immediately in saline and within 15-120 min (average of about 40 min) in 0.5% paraformaldehyde/2.5% glutaraldehyde. In 32 studied ganglia, 35% of 473 intrinsic cardiac neurons displayed striking pathological changes at the light and ultrastructural level. The other cells displayed normal morphology. The cytoplasm of 74% of the abnormal cells had one or more of three types of inclusions: (1) darkly stained lamellated inclusions (Type I), (2) membrane-bound whorls and parallel arrays of lightly stained membranes, as well as fine granular material (Type II), or (3) concentric layers of lightly stained membranes with a darker, granular core (Type III). Neurons with inclusions were markedly enlarged (66 x 54 microm vs. 40 x 34 microm for normal neurons) and displayed fewer dendrites. Some neurons contained electron lucent vacuoles indicative of degeneration while others showed frank degeneration, being fragmented, shrunken, and misshapen. Phagocytic cells containing lamellated inclusions and cellular debris were found in ganglia with abnormal neurons. Some axon terminals also displayed degenerative changes. The identification of pathological changes in the human intrinsic cardiac nervous system has implications with respect to the functional integrity of this final common regulator of cardiac function in disease states.

The ultrastructural effect and subcellular localization of mercuric chloride and methylmercuric chloride in insect cells (Aedes albopictusC6/36)

Abstract.The ultrastructural effects of mercuric chloride (Hg) and methylmercuric chloride (MeHg) were studied in theAedes albopictusC6/36 cell line. Both metal salts caused nuclear indentations, chromatin clumping and proliferation of the nucleolus. The mitochondria became pleomorphous. An increase of both free and membrane-bound ribosomes, swelling of the rough endoplasmic reticulum caused by accumulated protein and the appearance of well developed Golgi stacks all indicated the activation of protein synthesis. The activation was probably a cellular response to general stress, and the synthesized proteins may be members of the heat shock protein family. Apart from these common ultrastructural features, Hg-treated cells showed typical clusters of small electron-lucent vacuoles near the Golgi stacks. In cells exposed to MeHg, cytoplasmic tube-like structures were often observed and the disorganization of the organelles together with the appearance of blebs suggested disruption of the microtubules. Mercury accumulation was localized by an autometallographical silver staining technique both at the light and electron microscopic level; silver deposits were quantified by image analysis. For both Hg- and MeHg-treated cells, the degree of silver staining increased rapidly with increasing exposure time, but a considerable heterogeneity within the cell population was found. Lysosomes proved to be the major mercury storage sites in theAedescells and silver deposits could already be found after 30 min of Hg treatment. At sublethal concentrations, Hg inhibited the lysosomal marker enzyme acid phosphatase to some extent. For MeHg, no effect on this enzyme was found.

The ultrastructure and possible functions of nephrocytes in the trombiculid mite Hirsutiella zachvatkini (Acariformes: Trombiculidae)

The ultrastructure of nephrocytes during the post-larval development of the trombiculid mite Hirsutiella zachvatkini (Schluger) was investigated by means of transmission electron microscopy. Nephrocytes are situated either individually or in groups inside the haemocoelic space and are always ensheathed by a basal membrane. They contain numerous tubular elements, electron-dense inclusions and sometimes electron-lucent vacuoles. Invaginations of the plasma membrane, often in the form of labyrinthine channels, with coated pits of plasma membrane and characteristic slit diaphragms linking adjacent pedicels, typically occupy the peripheral parts of the cells. However, no distinct zonation of organelles in the nephrocytes of trombiculids was observed. The number of nephrocytes in the haemocoel and the intensity of development of their tubular elements and in particular of labyrinthine channels vary significantly during the ontogenesis of mites. The possible functions of nephrocytes and their structure and differentiation in comparison with other arachnids are discussed.

A fluid-phase endocytotic capacity and intracellular degradation of a foreign protein (horseradish peroxidase) by lysosomal cysteine proteinases in the rat junctional epithelium.

We investigated the co-localization of lysosomal cathepsins B, H and L, and horseradish peroxidase (HRP) in junctional epithelial (JE) cells both as a fluid-phase endocytotic marker to demonstrate the fluid-phase endocytotic capacity of JE cells, and to understand the morphological relationships of the endocytosed foreign substances to lysosomal cathepsins in these cells. The diaminobenzidine (DAB) histochemical and cytochemical methods and immunohistochemical avidin-biotin-peroxidase complex and immunocytochemical post-embedding colloidal gold methods were used. Under light microscopy, DAB reaction products based on HRP were found in JE but were rare or absent in the oral sulcular epithelium and oral epithelium. Immunolabeling for cathepsins B and H was found in the granular structures of the cells, but no cathepsin L was identified. With electron microscopy, DAB reaction products, which indicated both HRP and the azurophil granules of neutrophils, were endocytosed into JE cells. Using a post-embedding technique, gold particles indicating HRP were present on the plasma membrane of JE cells, at the periphery of electronlucent vacuoles, and in the electrondense granules. Gold particles indicating cathepsin B or H were found in the electrondense granules. With different sizes of colloidal golds, the co-localization of cathepsin B or H with HRP was indicated only in the electrondense portion of the larger vacuoles consisting of electronlucent and -dense parts. This study provided the first morphological data which indicate that JE has a fluid phase endocytotic capacity, and which suggest that the lysosomal cathepsins B and H are involved in the intracellular degradation of foreign substances invading through the gingival sulcus in JE cells.

Regulation of osteoclast function.

Regulation of osteoclast function.

Ultrastructural localization of basic proteins ofBlastocystis hominis

Basic proteins ofBlastocystis hominis were detected by the ammoniacal silver and ethanolic phosphotungstic acid techniques using electron microscopy. The central vacuole showed many silver grains when treated with ammoniacal silver and an increased electron density when treated with phosphotungstic acid. The intensity of positive reactions correlated with the electron density of the central vacuole, because cells having an electron-lucent central vacuole showed no silver grain deposits. Since it is known that the concentration of electron-dense materials in the central vacuole increases during log phase of growth, and then decreases in stationary phase, this organelle must accumulate basic proteins during cell growth.

Spermatogenesis in the endosymbiont-bearing bivalve Loripes lucinalis (Veneroida: Lucinidae).

In the endosymbiont-bearing bivalve Loripes lucinalis, spermatogenesis is similar to that described for numerous bivalve species and leads to the formation of an aquasperm. The head and midpiece measure 10.5 +/- 1.5 microns in length. The head is made up of a cylindrical nucleus slightly tapered apically and capped by a short conical acrosome. The nucleus lacks both an anterior and posterior nuclear invagination. The acrosome is 1.0 +/- 0.1 micron long and consists of an acrosomal cone containing a diffuse subacrosomal material and an apical electron-lucent vacuole. There is no axial rod. The midpiece is made up of four mitochondrial spheres that surround the distal and proximal centrioles. The base of the distal centriole is joined to the plasmic membrane by the pericentriolar complex made up of nine radial arms. A cytoplasmic collar is observed that sheaths the flagellum as it emerges from the distal centriole. The spermatozoa present in mature acini are grouped into characteristic rings, which may have a nutritive function, with the acrosome oriented toward the centre of these ring formations. Also present within the gonad are somatic cells that seem to play a nutritive role in relation to the germinal cells. These nutritive cells undergo a cycle of development and lysis that corresponds to the spermatogenic cycle of the bivalve. These cells are large and rich in glycogen and lipid inclusions. In-depth examination of nutritive cells and gametes reveals that the male gonad is devoid of microorganisms in either a vegetative or cryptic form, suggesting that a vertical transmission through paternal gonadal inheritance is a very unlikely means of symbiont transmission in L. lucinalis.

Morphological changes in the central vacuole ofBlastocystis hominis during in vitro culture

Morphological changes in the central vacuole during the growth in in vitro culture ofBlastocystis hominis were investigated by light and electron microscopy. Most cells in log phase and an early stationary phase showed a positive staining reaction in the central vacuole with PAS or Sudan black B stain, whereas cells in late stationary phase showed few positive reactions. Electron microscopic observations revealed that 95% ofB. hominis cells in log phase and 50% of cells in early stationary phase, had a substantial accumulation of electron-dense material in the central vacuole. In contrast, only 25% of the organisms in late stationary phase had an electron-dense central vacuole, while more than 50% of cells had an electron-lucent central vacuole. These results indicate thatB. hominis accumulated carbohydrates and lipids in the central vacuole during cell growth and that the organism probably consumed these metabolic substances during stationary growth. Therefore, it is strongly suggested that the central vacuole is an important organelle for storage of metabolic substances, such as carbohydrates and lipids, required for cell growth.

Persistent erythema and pruritus, with a confluent histiocytic skin infiltrate, following the use of a hydroxyethylstarch plasma expander

A 40-year-old woman developed a persistent pruritus with erythema of the head and upper trunk, which started within 2 weeks of a hydroxyethylstarch (HES) infusion and was still present 2 years later. Histological examination demonstrated a striking dermal infiltrate of KP1-positive foamy macrophages which had electron-lucent vacuoles. The timing of the onset, the body site distribution, the exclusion of other possibilities and an appropriate history of high dosage HES infusion, suggested that this was the cause of the eruption. Staining with periodic acid-Schiff (PAS) surprisingly was negative but we could not stain HES in vitro with this histochemical stain, and we postulate that the unique extent of the infiltrate and the negative PAS stain in our patient may indicate an impaired ability to degrade this chemical. This may be an important factor in the poorly understood pathogenesis of persistent pruritus induced by HES.

Vacuole-containing cells in the body cavity ofPhallmia mammillatalarvae before and after hatching

Summary The cells of the body cavity of Phallusia mammillata larvae were examined by transmission electron microscopy before hatching, after it and at the beginning of tail retraction. The results revealed the presence—both before and after hatching—of cells with small electron-lucent vacuoles, which might be the precursors of the monovacuolated cells observed at the beginning of tail retraction. These cells share morphological characteristics with the vanadium-accumulating signet ring cells of the blood of adults.

Wortmannin, a specific inhibitor of phosphatidylinositol-3 kinase, blocks osteoclastic bone resorption

The biological role of phosphatidylinositol (PI)-3 kinase was examined in osteoclast-like multinucleated cells (OCLs) formed in co-cultures of mouse osteoblastic cells and bone marrow cells. The expression of PI-3 kinase in OCLs was confirmed by Western blot analysis. Wortmannin (WT), a specific inhibitor of PI-3 kinase, inhibited PI-3 kinase activity in OCLs both in vitro and in vivo. WT also inhibited pit-forming activity on dentine slices and disrupted a ringed structure of F-actin-containing dots (an actin ring) in OCLs in a dose-dependent manner. The inhibitory profiles of WT for pit and actin ring formation were similar to that for PI-3 kinase activity in OCLs. Electron microscopic analysis revealed that OCLs treated with WT did not form ruffled borders. Instead, numerous electron lucent vacuoles of differing sizes were found throughout the cytoplasm. These results suggest that PI-3 kinase is important in osteoclastic bone resorption.

Atypical granules in the eyes of the white mutant of Drosophila melanogaster are lysosome-related organelles.

In the pigment cells of the white mutant of Drosophila melanogaster, as described earlier, two types of abnormal granules are found by conventional electron microscopy. However, both types of abnormal granules, in addition to those in pigment cell invaginations, are also present in the cytoplasm of the photoreceptor cells. Three enzymes (acid phosphatase, peroxidase, and tyrosinase) are localized within the eyes of wild type and white mutant Drosophila melanogaster by electron microscopy. Peroxidase activity is present in lamellar bodies close to the rhabdomeral microvilli of both fly types. However the organelles containing peroxidase activity are 6-fold more frequent in the wild type than in the mutant. Acid phosphatase is present in lamellar bodies between and at the bases of the rhabdomeral microvilli of the wild type, as well as in ommochrome granules of the photoreceptor cells. In the white mutant, however, acid phosphatase was located in electron lucent vacuoles in the cytoplasm of the receptor cells. These acid phosphatase-positive vacuoles also contained both types of abnormal granules. The latter result indicates that abnormal granules in the receptor cells originate from lysosomal degradation and that targeting of lysosomal enzymes is altered in the white mutant. Due to the tyrosinase activity in the hemolymph of flies, the extracellular spaces are electron dense after DOPA incubation. Since some abnormal granules within the photoreceptor cells are not surrounded by an extracellular space, they can be assumed to originate within the photoreceptor cells.

Fine structural aspects of the anterior necrotic zone in the leg bud of the chick embryo.

A fine structural study was undertaken on the area called the anterior necrotic zone of leg buds of the chick embryo at stages 22-29. Physiologically dying cells, which were distributed in the mesenchymal area, were characterized by increased electron density of the cytoplasm, appearance of electron lucent vacuoles and condensed chromatin pattern. Many of the dying cells seemed to be fragmented into several pieces of cell debris during the necrotic process. Two kinds of cells were observed to take up the necrotic cells; one type, large cells 15 microns or more in size and irregular in shape, which had lysosomes, extended microvillus-like cytoplasmic projections and pseudopods, took up pieces of necrotic cells by phagocytosis and stored them in acid phosphate-negative and -positive vacuoles. The other type was several microns in diameter with numerous free ribosomes and stored a small number of vacuoles containing cell debris. At stage 29, profiles of necrotic cells lying free in the mesenchymal area, and cells containing one or two vacuoles filled with cell debris decreased in number as well as large cells containing numerous vacuoles, whose contents seemed to be degraded. When pieces of the anterior portion of chick leg buds at stage 24 were organ-cultured for 3 days after glutaraldehyde-fixed mouse red blood cells were injected, the red blood cells were taken up into cells in the mesenchymal area. Formation of large cells containing numerous vacuoles stuffed with red blood cells was rare. These findings may imply that most necrotic cells in the anterior necrotic zone are disposed of by macrophages and that some are endocytosed by mesenchymal cells.

Isolated fish hepatocytes as a tool in aquatic toxicology: sublethal effects of dinitro- o-cresol and 2,4-dichlorophenol

In order to evaluate the substance-specificity of ultrastructural criteria in isolated fish hepatocytes as endpoints in toxicological studies, liver cells isolated from rainbow trout ( Oncorhynchus mykiss) by collagenase perfusion were exposed in vitro to sublethal concentrations of dinitro-o-cresol (DNOC: 0, 0.7, 7.2, 72.2 μmol/1) and 2,4-dichlorophenol (2,4-DCP: 0, 6.1, 61.3, 613 nmol/1) for 1–5 days. With either toxicant, earliest cytological modifications could be detected after 1 day at the lowest test concentrations. Ultrastructural analysis documents a complex, substance-specific combination of dose- and time-dependent effects. Evaluation of single cytological alterations induced by DNOC and 2,4-DCP reveals that each cytopathological reaction consists of both unspecific and substance-specific changes. Unspecific symptoms comprise disturbance of intracellular compartmentation, condensation, marginalization and redistribution of heterochromatin, reduction and reorganization of RER cisternae, proliferation of lysosomes, glycogenosomes, myelinated bodies and large electron-lucent vacuoles with heterogeneous contents, as well as glycogen depletion. Transformation of Golgi cisternae into concentric membrane whorls is specific to intoxication by DNOC, whereas the reaction to 2,4-DCP is characterized by unusual heterochromatin distribution patterns and formation of mitochondrion aggregations. Due to their general character, unspecific changes are recommended as rapid biomarkers of contamination by chemicals, whereas specific changes might be useful for the differential diagnosis of the toxicants.

Fine structure of the neurohemal sinus gland of the shore crab, Carcinus maenas, and immuno-electron-microscopic identification of neurosecretory endings according to their neuropeptide contents.

The sinus gland of the shore crab, Carcinus maenas, is a compact assembly of interdigitating neurosecretory axon endings abutting upon the thin basal lamina of a central hemolymph lacuna. Four types of axon endings are distinguishable by the size distribution, shape, electron density and core structure of their neurosecretory granules. One additional type of axon ending is characterized by electron-lucent vacuoles and vesicles. The axon profiles are surrounded by astrocyte-like glial cells. Various fixations followed by epoxy- or Lowicryl-embedding were compared in order to optimize the preservation of the fine structure of the granule types and the antigenicity of their peptide hormone contents. By use of specific rabbit antisera, the crustacean hyperglycemic, molt-inhibiting, pigment-dispersing, and red-pigment-concentrating hormones were assigned to the four distinct granule types which showed no overlap of immunostaining. Epi-polarization microscopy and ultrathin section analysis of immunogold-stained Lowicryl-embedded specimens revealed that immunoreactivity to Leu-enkephalin and proctolin is co-localized with molt-inhibiting hormone immunoreactivity in the same type of granule. The size and core structure of the immunocytochemically identified granule types vary little with the different pretreatments but, in some cases, to a statistically significant extent. The present results are compared with those from earlier studies of sinus glands in different crustaceans. The methods of granule identification used in this study supplement the classical approach in granule typing; they are easier to perform and more reliable for the analysis of release phenomena in identified secretory neurons supplying the neurohemal sinus gland.

Evidence for a pathway of distal screening pigment granules across the basement membrane of the crayfish photoreceptor.

The distal pigment cells of Orconectes limosus contain two layers of large electron lucent vacuoles that are separated by layers of small right-angled platelets adjacent to the crystalline cones. The crystalline cones of the dioptric apparatus of this species have evaginations into the distal pigment cell cytoplasm. In photoreceptors of Orconectes limosus and Procambarus clarkii a dark pigment accumulation site was detected just distal to the basement membrane at the edges of each retina. These pigment accumulations occurred independent of the state of light adaptation. Ultrastructurally the pigment granules at this accumulation site resemble distal screening pigment granules according to their size (up to 1.2 microns in diameter) and fibrous structure. Distal screening pigment granules were also found in tube-like cell processes or extracellularly within and proximal to the retinal basement membrane, indicating pigment transport to and across the basement membrane. Proximal to the basement membrane screening pigment granules were also observed disintegrated to a gravel-like electron dense material in widely branched cells. Evidence was found that an electron dense material, probably resulting from disintegrating screening pigment granules, was incorporated in the integument of the eyestalk. Four hours after injection of gold particles into the eye stalk distal to the retina they were detected inside and proximal to the retinal basement membrane.

Morphological evidence for a single bacterial etiology in Texas necotizing hepatcpanceatitis in Penaeus vannamei (Crustacea. Decapoda)

Morphological evidence for a single bacterial etiology in Texas necotizing hepatcpanceatitis in Penaeus vannamei (Crustacea. Decapoda)