Multivesiculate Bodies Research Articles

Photosynthetic, antioxidative, molecular and ultrastructural responses of young cacao plants to Cd toxicity in the soil

Cadmium (Cd) is a highly toxic metal for plants, even at low concentrations in the soil. The annual production of world cocoa beans is approximately 4 million tons. Most of these fermented and dried beans are used in the manufacture of chocolate. Recent work has shown that the concentration of Cd in these beans has exceeded the critical level (0.6mgkg−1 DM). The objective of this study was to evaluate the toxicity of Cd in young plants of CCN 51 cacao genotype grown in soil with different concentrations of Cd (0, 0.05 and 0.1gkg−1 soil) through photosynthetic, antioxidative, molecular and ultrastructural changes. The increase of Cd concentration in the soil altered mineral nutrient absorption by competition or synergism, changed photosynthetic activity caused by reduction in chloroplastidic pigment content and damage to the photosynthetic machinery evidenced by the Fv/Fm ratio and expression of the psbA gene and increased GPX activity in the root and SOD in leaves. Additionally, ultrastructural alterations in roots and leaves were also evidenced with the increase of the concentration of Cd in the soil, whose toxicity caused rupture of biomembranes in root and leaf cells, reduction of the number of starch grains in foliar cells, increase of plastoglobules in chloroplasts and presence of multivesiculated bodies in root cells. It was concluded, therefore, that soil Cd toxicity caused damage to the photosynthetic machinery, antioxidative metabolism, gene expression and irreversible damage to root cells ultrastructure of CCN 51 cocoa plants, whose damage intensity depended on the exposure time to the metal.

Up-regulation of Vps4A promotes neuronal apoptosis after intracerebral hemorrhage in adult rats

Vps4, vacuolar protein sorting 4, belongs to ATPases Associated with diverse cellular Activities (AAA) protein family which is made up of Vps4A and Vps4B. Previous studies demonstrated that Vps4A plays vital roles in diverse aspects such as virus budding, the efficient transport of H-Ras to the PM (plasma membrane) and the involvement in the MVB (multivesiculate bodies) pathway. Interestingly, Vps4A is also expressed in the brain. However, the distribution and function of Vps4A in ICH diseases remain unclear. In this study, we show that Vps4A may be involved in neuronal apoptosis during pathophysiological processes of intracerebral hemorrhage (ICH). Based on the results of Western blot and immunohistochemistry, we found a remarkable up-regulation of Vps4A expression surrounding the hematoma after ICH. Double labeled immunofluorescence showed that Vps4A was co-expressed with NeuN but rarely with astrocytes and microglia. Morever, we detected that neuronal apoptosis marker active caspase-3had co-localizations with Vps4A. Additionaly, Vps4A knockdown in vitro specifically leads to decreasing neuronal apoptosis coupled with increased Akt phosphorylation. All datas suggested that Vps4A was involved in promoting neuronal apoptosis via inhibiting Akt phosphorylation after ICH.

Influenza virus budding does not require a functional AAA+ ATPase, VPS4

The process of budding of many enveloped viruses utilizes the cellular ESCRT (endosomal sorting complex required for transport) machinery, that is normally involved in the formation of luminal vesicles of endosomal multivesiculate bodies (MVB). A late step in the MVB pathway involves the recruitment of VPS4, an AAA+ ATPase, to the ESCRT complexes. Our earlier work had shown that the formation of influenza virus-like particles was not inhibited by dominant negative VPS4A. However, it was not known if there was a role of VPS4 and the ESCRT pathway in influenza virus particle budding and this needed to be investigated. It was found that neither siRNA knockdown of VPS4A and VPS4B expression nor the use of cell lines that inducibly express VPS4A or VPS4B dominant negative mutants, inhibited influenza virus budding. In contrast, and in keeping with more recent data, vesicular stomatitis virus budding was diminished by VPS4 dysfunction.

The Vps27p Hse1p complex binds ubiquitin and mediates endosomal protein sorting.

Membrane proteins that are degraded in the vacuole of Saccharomyces cerevisiae are sorted into discrete intralumenal vesicles, analogous to the internal membranes of multi-vesiculated bodies (MVBs). Recently, it has shown that the attachment of ubiquitin (Ub) mediates sorting into lumenal membranes. We describe a complex of Vps27p and Hse1p that localizes to endosomal compartments and is required for the recycling of Golgi proteins, formation of lumenal membranes and sorting of ubiquitinated proteins into those membranes. The Vps27p-Hse1p complex binds to Ub and requires multiple Ub Interaction Motifs (UIMs). Mutation of these motifs results in specific defects in the sorting of ubiquitinated proteins into the vacuolar lumen. However, the recycling of Golgi proteins and the generation of lumenal membranes proceeds normally in Delta UIM mutants. These data support a model in which the Vps27p-Hse1p complex has multiple functions at the endosome, one of which is as a sorting receptor for ubiquitinated membrane proteins destined for degradation.

Immunoelectron microscopic investigations of patching, capping, endocytotic and shedding processes in T and B lymphocytes

Lymphocytes were isolated from the blood of healthy juvenile test persons by the FICOLL method. Subsequently, CD4-, CD8-, and CD19-positive cells were obtained by the use of magnetic beads. The sandwich technique and gold labelling method (preembedding) served for the demonstration of receptors in the electron microscope. The gold-labelled receptors were primarily endocytosed via smooth-walled micropinocytotic processes, less frequently by coated pits/vesicles. The endocytosis cycle lasted only as far as the level of multivesiculated bodies. Lysosomes and structures of the Golgi apparatus were free from gold particles. It was surprising that after activation the capping phenomena were not associated with increased endocytotic activity. The inner surface of the membrane of endocytotic vesicles underneath the cap does not explain modulation or turnover of the receptor under these conditions, not even in view of a fast endocytotic cycle. Another possibility of a membrane turnover is the "shedding" process. We were indeed able to demonstrate gold labelling and surface coat-like material in the extracellular matrix.

Fine Structure of the Mature Plasmodium of Intoshia variabili (Phylum Orthonectida), a Parasite of the Platyhelminth Macrorhynchus crocea

AbstractThe fine structure of the mature plasmodia of Intoshia variabili, found in the kalyptorhynch flatworm Macrorhynchus crocea, was studied with transmission electron microscopy. As the Plasmodium matures it forms long finger‐like extensions inside the host. These extensions eventually take the place of the genital system of the host turbellarian. The mature Plasmodium is separated from the host tissues by a continuous membrane. The entire Plasmodium is surrounded by modified host cells, which are thin and overlapping. These cells also have a rather homogenous cytoplasm with limited endoplasmic reticulum. The surface of the Plasmodium is mostly smooth and has numerous small extensions. Within the cytoplasm of the Plasmodium, different stages of development of both males and females could be observed. The cytoplasm also contains numerous small nuclei with distinct nucleoli. Scattered throughout the cytoplasm, in addition to nuclei, are mitochondria, various granules, multivesiculate bodies with lamellar structures, and a large number of small vesicles. The nature of the formation of the Plasmodium is discussed. © 1998 The Royal Swedish Academy of Sciences. Published by Elsevier Science Ltd. All rights reserved

Ultrastructure of gametes after intracytoplasmic sperm injection.

The ultrastructure of 18 metaphase-II oocytes after intracytoplasmic sperm injection (ICSI) was analysed from 30 min to 8 h following the microinjection. Three control metaphase-II oocytes were not injected. The 21 oocytes were embedded individually and examined in serial pole-to-pole sections. Seventeen oocytes had been matured in vitro from the germinal vesicle stage and showed ultrastructural alterations due to long-term culture. In microinjected oocytes, membrane-bound vacuoles and oolemma inclusions were found at the injection site. The ICSI oocytes showed evidence of plasma-membrane damage and increased oocyte exchange processes with residual and multivesiculated bodies. Gamete membrane fusion was not observed. Acrosome reaction was observed within the ooplasm 15 min after ICSI. Sperm elements were not incorporated into any oocyte vacuole. Onset of oocyte activation was associated with sperm chromatin decondensation. Cortical granules were released at 60 min and two pronuclei were formed at 6 h after ICSI. The male and female pronucleus formation was asynchronous. The findings suggest specific ICSI-related morphological features of early fertilization.

Fine structure of the developing oocytes in adult Argas (Persicargas) arboreus (Ixodoidea: Argasidae).

The structure of the developing oocytes in the ovary of unfed and fed female Argas (Persicargas) arboreus is described as seen by scanning (SEM) and transmission (TEM) electron microscopy. The unfed female ovary contains small oocytes protruding onto the surface and its epithelium consists of interstitial cells, oogonia and young oocytes. Feeding initiates oocyte growth through the previtellogenic and vitellogenic phases of development. These phases can be observed by SEM in the same ovary. The surface of isolated, growing oocytes is covered by microvilli which closely contact the basal lamina investing the ovarian epithelium and contains a shallow, circular area with cytoplasmic projections and a deep pit, or micropyle, at the epithelium side. In more advanced oocytes the shell is deposited between microvilli and later completely covers the surface. Transmission EM of growing oocytes in the previtellogenic phase reveals nuclear and nucleolar activity in the emission of dense granules passing into the cytoplasm and the formation of surface microvilli. The cell cytoplasm is rich in free ribosomes and polysomes and contains several dictyosomes associated with dense vesicles and mitochondria which undergo morphogenic changes as growth proceeds. Membrane-limited multivesiculate bodies, probably originating from modified mitochondria, dictyosomes and ribosomal aggregates, are also observed. Rough endoplasmic reticulum is in the form of annulate lamellae. During vitellogenesis, proteinaceous yolk bodies are formed by both endogenous and exogenous sources. The former is involved in the formation of multivesicular bodies which become primary yolk bodies, whereas the latter process involves internalization from the haemolymph through micropinocytosis in pits, vesicles and reservoirs. These fuse with the primary yolk bodies forming large yolk spheres. Glycogen and lipid inclusions are found in the cytoplasm between the yolk spheres.

Ultrastructural changes of Chaetomorpha brachygona growing in metal environment.

The structural changes of the filamentous green alga C. brachygona growing in iron-ore tailing which contained high levels of manganese, cadmium, chromium, copper, iron, lead and zinc were investigated with the electron microscope. The most conspicuous ultrastructural change of the algal cells was the substantial increase in the thickness of the side wall and the cross wall between adjacent cells, which could trapped large amounts of metal precipitates. The well lamellated outer portions of the thickened side wall probably functioned in preventing rapid movement of the metal precipitates through the wall into the cell lumen. There was an increase in the number as well as the size of the lipid bodies in the metal-tolerant algal cells. Reduction of the size of the chloroplasts which showed a reduced internal organization was also observed. In addition, the presence of multivesiculate bodies in the cytoplasm which were not detected in normal algal cells, served as reservoir for the sequenstration of metal ions in the cytoplasm. Deposition of metal precipitates was not detected in the chloroplast, mitochondria and nucleus of the cell however. The metal tolerance exhibited by C. brachygona was therefore attributed to these ultrastructural changes of the algal cells, which enabled the algal filaments to survive and maintain growth in the metal-riched environment.

Colchicine-induced tubular, vesicular and cisternal organelle aggregates in absorptive cells of the small intestine of the rat. II.--Endocytosis studies.

Administration of the antimicrotubular agent colchicine to adult rats (0.5 mg/100 g of body weight for 6 hr) induces formation of extended aggregates of tubular, vesicular, and cisternal organelles in the absorptive cells of the small intestine. The phosphatase reaction pattern (thiamine pyrophosphatase, acid phosphatase, acid trimetaphosphatase) suggests that the majority of them belongs to the lysosomal system (Ellinger and Pavelka, 1984). The present study extends these findings and examines the uptake and fate of intravenously injected horseradish peroxidase (HRP) at the basal and lateral cell surfaces and of intraluminally applied HRP at the apical cell surface. HRP, applied to control animals and animals pretreated with colchicine, was internalized at both apical and basolateral cell surfaces of the absorptive cells, and delivered into endosome-like vesicles, multivesiculated bodies (mvbs), dense bodies (dbs), and in several instances into Golgi cisternae. Following intraluminal application, evidence was obtained for the transport of HRP across the cell; in contrast, intravenously applied HRP was never detected at the apical cell surface. Colchicine pretreatment did not stop the uptake of HRP, which was rapidly sequestered to the clustered tubules, vesicles, and cisternae, as well as to the mvbs and dbs. After longer intervals, the portion of HRP-reactive tubules, vesicles, and cisternae within the clusters increased: 60 min after HRP-administration all of them contained HRP-activity. These results indicate that the colchicine-induced clustered organelles are recipients of endocytic materials internalized at the apical as well as at the basolateral cell surface.

Electron microscopic studies on IgA nephropathy.

We carried out electron microscopic studies on renal tissues from 9 patients with IgA nephropathy. Electron dense deposits were present in the mesangial area in all cases, subendothelial deposits in 4, and subepithelial deposits in only one. In basement membrane, segmental swelling and rarefaction of basement membrane substance were observed. In some cases the degenerated basement membrane substance protruded through the dilated endothelial fenestration into capillary lumina. Focal splitting, attenuation, mouse eaten appearance, and herniation of basement membrane were seen in a high incidence. Mesangial cells possessed well developed rough endoplasmic reticulums and polysomes. In the peripheral areas of mesangial cell cytoplasm, there was accumulation of electron dense substance and this was occasionally continuously present in the mesangial matrix. There was segmental swelling of endothelial cell cytoplasm, resulting in loss of fenestration. Epithelial cells had well developed rough endoplasmic reticulums and polysomes. Segmental foot process fusion was seen, and these processes, regardless of fusion, became electron denser in the area close to the basement membrane. Multivesiculated bodies were present in all cases in the epithelial cells and in 7 cases in the endothelial cells. Spherical microparticles were present in 3 cases in the urinary space or between the basement membrane and the epithelial cells.

Rhabdome-bearing cells within the corpus cardiacum of Melanogryllus desertus (Orthoptera, Gryllidae)

Cells with rhabdomeric structures characteristic of photoreceptor cells occur within the corpus cardiacum of Melanogryllus desertus. These cells are few in number and are larger than intrinsic secretory cells. Each has numerous rhabdomeric microvilli forming a rhabdome within the cell. The arrangement of large mitochondria under the rhabdomeric microvilli around the dilated endoplasmic reticulum membranes and multivesiculated bodies is distinctive. Rhabdome cells have synaptic contacts with the numerous neurosecretory axons that surround them. Other neurosecretory axons come in close contact with the membranes of microvilli.

The aging vestibular hair cell

The ultrastructure of aging vestibular hair cells of the guinea pig can be abnormal even though they appear morphologically normal by light microscopy. In only a few of the hair cells studied did nonspecific changes occur. In general, the age-related changes involved specific structures in the hair cells: aggregations of lipofuscin pigments, multivesiculated bodies, disintegration of the cuticular plate, and rod-shaped inclusions from the cuticle area into the hair cell. Furthermore, sensory hairs are generally present when the cuticular plate shows an advanced disintegration. Nerve calyces can be ultrastructurally changed without morphologic changes in the adjacent hair cell. Type I hair cells show age-related morphologic changes more often than do type II hair cells.

The effect of heavy metals on the cytoplasmic fine structure ofSkeletonema costatum (Bacillariophyta)

The marine diatomSkeletonema costatum (Greville) Cleve was grown in batch culture in the presence of mercury (6.5 μg 1−1), cadmium (50 μg 1−1) and zinc (265 μg 1−1). Alterations in cytoplasmic morphology were observed in cells treated with these sublethal concentrations of the metals. Swollen organelles, dilated membranes and vacuolated cytoplasm indicated plasma membrane damage and subsequent osmotic disorganization. Electron-dense inclusions in vesicles, multivesiculate bodies and cytoplasmic “tubules” apparently derived from the Golgi body are interpreted as mechanisms of sequestering the metals. Mercury and zinc had a marked effect on the fine structure whereas treatment with cadium resulted in little to no cytoplasmic damage.

Scanning and transmission electron microscopy of the ependyma of the fourth ventricle in the monkey brain.

The ependymal structure of the fourth ventricle floor of adult rhesus monkeys was investigated using both scanning and transmission electron microscopy. Each half of the floor exhibited dense ciliation which decreased towards the median sulcus. The area postrema and the facial colliculus lacked cilia and were covered with non-ciliated ependymal cells. Along the entire length of the median sulcus supraependymal globular protrusions were seen. Over the rostral and middle thirds of the sulcus these protrusions exhibited interwoven ridges. At the former situation they were closely approximated but at the latter they were on the surface of non-ciliated nodules which were lying amongst sparsely ciliated cells. In the caudal third these protrusions presented smoother surfaces without any structural details over them. Transmission electron microscopy through the upper two thirds of the median sulcus revealed these surface protrusions to be non-ciliated cells having long and profuse network of interwoven microvilli over their luminal surface. These cell bodies exhibited well-defined supranuclear Golgi complexes, vesiculated rough and smooth surfaced endoplasmic reticulum, free ribosomes and polyribosomes, few electron dense granules, network of microtubules and light and dense multivesiculated bodies. These features suggested a secretory and/or absorptive role, rather that autophagy to these surface specialization.

Electron microscope study of the epimastigotes of a fish trypanosome, Trypanosoma cobitis, in culture.

The ultrastructure of the epimastigotes of the fish trypanosome, Trypanosoma cobitis, cultured in 4N blood agar has been examined. The main features of interest are the presence of microorganisms, pigment bodies, long, curved and circular mitochondrial cristae, a cytopharyngeal complex and multivesiculate bodies.

Ultrastructure of the Epimastigotes of the Fish Trypanosome Trypanosoma cobitis Mitrophanow 1883, in the Crop of the Leech Vector, Hemiclepsis marginata

The ultrastructure of the epimastigote of Trypanosoma cobitis in the crop of the leech vector Hemiclepsis marginata is described. Microorganisms, closely resembling endosymbiotic bacteria, seen in monoxenous trypanosomatids, were present in the cytoplasm. Also, within the cytoplasm were ribosomes, sparse endoplasmic reticulum, a Golgi apparatus, and a wide variety of other inclusions, in particular multivesiculate bodies which appeared to contain waste material. The well-developed branched mitrochondrion had platelike cristae and an expansion near the nucleus which contained the kinetoplast DNA. A cytostomal opening situated near the flagellar pocket leads to a cytopharynx surrounded by five microtubules.

Spermiogenesis in Nautilus pompilius. II. Sertoli cell-spermatid junctional complexes.

A "ball and socket-like" junction between branches of the Sertoli cells and the developing spermatids is described. A cytoplasmic extension of the Sertoli cell fits into a pocket in the spermatid and has a constricted neck region. Frequently there are large multivesiculate bodies in the Sertoli cell extension and small vesicles frequently appear in the spermatid cytoplasm, in the area of the "ball and socket-like" junction. This suggests the possibility that there may be communication of materials between the two cells. The possible function of the junctions is discussed and it is concluded that they more likely have a role in nutrition than in coordination.

Cellular responses to surface binding and internalization of Concanavalin A: An electron microscopic investigation on the problem of membrane cycling

Monolayer cultures of normal and diethylnitrosamine-transformed rat liver cells were labeled in situ with Con A-HRP or ferritin-conjugated Con A. Ligand-induced redistribution with simultaneous internalization of labeled membrane areas occurred in normal as well as in transformed cells when they were reincubated with PBS at 37 degrees C for different periods of time (from 5 min up to 3 hrs). Compared to normal cells, these afore mentioned processes were accelerated in transformed cells. Internalization in normal and transformed cells resulted in a recycling of labeled plasma membrane areas in the Golgi region with the label being finally accumulated in elements which correspond mostly, but not exclusively, to GERL. Then formation of phagolysosomes and multivesiculated bodies occurred whose labeled content was exocytized after fusion with the plasma membrane. This suggested that the internalized plasma membrane areas were at least partly degraded. The relabeling of some parts of the plasma membrane by extruded lysosomal content indicates that at least some Con A molecules are still biological active. Membrane internalization by endocytosis after binding of Con A obviously causes an increased of membrane biogenesis and exocytosis, thus compensating for membrane removal. This is suggested by the vacuolization and enlargement of unlabeled (not in recycling involved) Golgi apparatus. It may indicate a differential functional role of the Golgi apparatus in membrane turnover in the same cell. The fusion of phagolysosomes with the plasma membrane and the insertion of phagolysosomal membrane into the plasma membrane might be another compensatory mechanism.

Fine structure of two types of pleuropodia in Diploptera punctata (Dictyoptera: Blaberidae) with observations on their permeability

Fine structural analysis of the 2 types of pleuropodia of embryos of the viviparous cockroach, Diploptera punctata (Eschscholtz) reveals that both organs have vast plasma membrane-mitochondrial complexes, but on opposite surfaces of their cells. The appendage-like pleuropodia on the 1st abdominal segment exhibit mitochondria within microvilli at the outer cell surfaces of the bulb and extension which comprise the bulk of the organ. Lateral surfaces are interdigitated and mitochondria-rich, but the inner surfaces are relatively free of mitochondria and infoldings. The atypical cuticle of the bulb is permeable to several vital dyes but less so than the extension, presumably because of a visibly different cuticle. The second type of pleuropodia, called adenopodia, are of the semi-invaginated type, located on the 9th abdominal segment. These organs have mitochondria within deep infoldings of the inner plasma membrane and along the lateral cell surfaces, but not in the microvilli and infoldings at the outer surfaces. Apical pinocytotic vesicles appear to contribute to the formation of multivesiculate bodies which occur in channels between tightly packed microtubules. It is unlikely that these vesicles mediate the uptake of large molecules from the extra-embryonic fluid because vital dyes do not penetrate the cuticle of the adenopodia. However, silver nitrate reacts strongly at its surface, suggesting the presence of and permeability to chloride. Since the structure of these organs is similar to that of ion and fluid transport tissues, such as malpighian tubules and anal papillae, it is proposed that their function as osmoregulators of embryonic hemolymph and extracellular fluid be investigated.