Differential immunocytochemical staining for glial fibrillary acidic (GFA) protein,S-100 protein and glutamine synthetase in the rat subcommissural organ,nonspecialized ventricular ependyma and adjacent neuropil

Summary Antibodies raised against glial fibrillary acidic protein (GFA), S-100 protein (S100) and glutamine synthetase (GS) are currently used as glial markers. The distribution of GFA, S100 and GS in the ependyma of the rat subcommissural organ (SCO), as well as in the adjacent nonspecialized ventricular ependyma and neuropil of the periaqueductal grey matter, was studied by use of the immunocytochemical peroxidase-antiperoxidase technique. In the neuropil, GFA, S100 and GS were found in glial elements, i.e., in fibrous (GFA, S100) and protoplasmic astrocytes (S100, GS). The presence of S100 in the majority of the ventricular ependymal cells and tanycytes, and the presence of GFA in a limited number of ventricular ependymal cells and tanycytes confirm the glial nature of these cells. The absence of S100, GFA and GS from the ependymocytes of the SCO, which are considered to be modified ependymal cells, suggests either a non-astrocytic lineage of these cells or an extreme specialization of the SCO-cells as glycoprotein-synthesizing and secreting elements, a process that may have led to the disappearance of the glial markers.     

Combined immunostaining of neurofilaments, neuron specific enolase, GFAP and S-100. A possible means for assessing the morphological and functional status of the enteric nervous system

Neurofilaments, part of the cytoskeletal network, and neuron specific enolase, a major enzyme in glycolysis, are both present in central and peripheral neurons. Glial fibrillary acidic protein and S-100, on the other hand, are soluble proteins which are found exclusively in the supportive cells of the nervous system, i.e. the glial cells. Examination was made, using immunocytochemistry, of all main areas of the gastrointestinal tract of three mammalian species, rat, pig and man. By applying serial tissue sectioning, it was possible to study the relative occurrences of the two neuronal markers in the same cell bodies and to examine the relationships of the neurons with the glial cells as revealed by the antibodies to glial fibrillary acidic protein and S-100. Both neurofilaments and neuron specific enolase were localised to an extensive system of enteric nerves, with the level of neuron specific enolase-immunoreactivity showing greater variability than that observed using antibodies to neurofilaments. Comparison of the occurrence of neuron specific enolase and neurofilament immunoreactivity in serially sectioned neuronal cell bodies revealed that a minor population stained only with antibodies to neurofilaments. The equivocal or absent neuron specific enolase-immunoreactivity in some perikarya may reflect variations in functional status within the nervous system. Glial fibrillary acidic protein- and S-100-immunoreactivities were confined to glial cells which, in this normal tissue, were always in close association with the neurons.(ABSTRACT TRUNCATED AT 250 WORDS)     

In vivo and in vitro differentiation of neurons and astrocytes in the rat embryo. Immunofluorescence study with neurofilament and glial filament antisera

Antisera raised against neurofilament (NF) peptides and glial fibrillary acidic protein (GFA) (subunit of glial filaments) have been used to identify neurons and astrocytes in order to study their development and differentiation in rat embryo. In vivo observations showed that NF-positive cells first appeared in 12-day-old embryos, whereas GFA-positive cells appeared in brain and spinal cord on the 18th day. In vitro observations showed that NF-positive cells could be obtained only in cultures from 12-day embryos onwards. The further differentiation of neurons involved neurite elongation, aggregation of cell bodies to form islets, and emergence of very brightly staining prominent neurons with large cell bodies and long neurites which took part in complicate pattern formation. GFA-positive cells appeared in vitro on the 16th day and they could be observed even in cultures obtained from 10-day-old embryos. As the culture aged, the GFA staining became highly fibrillary. There was no physical interaction between neuronal and glial processes.     

Morphogenesis and proliferation of the larval brain glia in Drosophila

Glial cells subserve a number of essential functions during development and function of the Drosophila brain, including the control of neuroblast proliferation, neuronal positioning and axonal pathfinding. Three major classes of glial cells have been identified. Surface glia surround the brain externally. Neuropile glia ensheath the neuropile and form septa within the neuropile that define distinct neuropile compartments. Cortex glia form a scaffold around neuronal cell bodies in the cortex. In this paper we have used global glial markers and GFP-labeled clones to describe the morphology, development and proliferation pattern of the three types of glial cells in the larval brain. We show that both surface glia and cortex glia contribute to the glial layer surrounding the brain. Cortex glia also form a significant part of the glial layer surrounding the neuropile. Glial cell numbers increase slowly during the first half of larval development but show a rapid incline in the third larval instar. This increase results from mitosis of differentiated glia, but, more significantly, from the proliferation of neuroblasts.     

An Ultrastructural Study of Cell-to-Cell Membrane Interaction at Early Stages of Postnatal Ontogenesis of Cortical Brain Neurons in White Rats

In early postnatal ontogenesis of cerebral cortex (visual area) of the white rat, a wide distribution of different types of membrane contacts have been found between developing nervous cells and their processes. The following types of contacts were observed: 1. Penetration of thin filopodia into specialized invaginations having all the features of coated vesicles; 2. Contacts of filopodia with thickened surface membrane; 3. Contacts of opposit filopodia; 4. Contacts of membranes with reciprocal invaginations alternating with filopodia or surface blebs.
These types of membrane interaction were regularly distributed along the surface of cells and their processes, and were situated in close approximation to typical tight junctions and other adhesional complexes. As a rule, filopodia were components of axon branches, and almost all invaginations were situated on plasma membranes of cell bodies or on dendrites, although sometimes there were invaginations on axon profiles and filopodia on dendrites.
It is suggested that the distribution and structural specialization of these membrane contacts reflect their participation in the process of programmed cell-to-cell recognition that precedes the formation of synaptic contact. Other reports and the current data reveal the special morphogenetic role of membrane communication in the formation and stability of integrative cell systems.     

Glial cells phagocytose neuronal debris during the metamorphosis of the central nervous system in Drosophila melanogaster

 Using electron microscopy we demonstrate that degenerating neurons and cellular debris resulting from neuronal reorganization are phagocytosed by glial cells in the brain and nerve cord of the fruitfly Drosophila melanogaster during the first few hours following pupariation. At this stage several classes of glial cells appear to be engaged in intense phagocytosis. In the cell body rind, neuronal cell bodies are engulfed and phagocytosed by the same glial cells that enwrap healthy neurons in this region. In the neuropil, cellular debris in tracts and synaptic centres resulting from metamorphic re-differentiation of larval neurons is phagocytosed by neuropil-associated glial cells. Phagocytic glial cells are hypertrophied, produce large amounts of lysosome-like bodies and contain a large number of mitochondria, condensed chromatin bodies, membranes and other remains from neuronal degeneration in phagosomes. Received: 23 January 1996 / Accepted in revised form: 21 May 1996     

Some aspects of the structural organization of the arthropod ganglia

Summary This paper deals with the fine structure of the abdominal ganglia of several species of arthropods belonging to the classes Arachnida, Crustacea, Myriapoda and Insecta. The tissues were fixed in osmium tetroxide and embedded in n-butyl methacrylate or fixed in potasium permanganate and embedded in a mixture of X 133/2097 and Araldite.A comparative study was made in order to discriminate between those structural characteristics of the nervous system appearing only in determined taxonomic groups and those belonging to a fundamental plan common to the whole Phylum. This work covers the morphology of neurons, glial cells, neuropilic nerve fibers and neuronal connections.Most arthropod neurons are pear-shaped with only one prolongation and the nucleus is located in the center of the soma, enveloped by two membranes showing numerous pores. Cisternae of the ER have frequently been observed in continuity with this nuclear envelope. After osmic fixation the nuclear content appears to consist of small dense granules distributed at random in the nucleoplasm. In addition to these small perticles there are, in some species, large chromatin blocks. The use of Permanganate as fixative introduces important changes in the nuclear aspect; most of the nuclei look washed and the nuclear content acquires an homogeneous appearance.The cytoplasm of the neurons contains a complex system of internal membranes consisting of cisternae and tubuli of the ER system, lamellae of the Golgi complex and invaginations of the plasma membrane. In most species the elements of the ER system are distributed at random in the cytoplasm but in the neurons of Bothriurus bonariensis there are parallel aggregations of membranes similar to the Nissl bodies found in vertebrates.It was found in some of the species studied (Armadillidium vulgare and Lithobius Sp.) that the internal membrane system of the nerve cells is mainly represented by Golgi elements while the ER system seems to be poorly developed.Besides the membranous components, the neuronal cytoplasm contains mitochondria, multivesicular bodies and dense granules of neurosecretory material.Neuroglial cells are mainly characterized by their nuclear structure. After the action of osmium tetroxide, glial nuclei show irregular masses of chromatin inmersed in a nucleoplasm of low electron density. In permanganate fixed material these chromatin blocks appear as blank spaces.In the cytoplasm of these cells there are mitochondria, membranes pertaining to the ER system and elements of the Golgi complex but in some of the species studied gliofibrils and granules of pigment were found.Three main types of neuroglial cells have been recognized in an arthropod ganglia. These are: subcapsular glial cells, neuron satellites and nerve fiber satellites.The neuropile occupies the central region of the ganglion and consists of a great number of nerve fibers intermingled with glial processes. The neuropilic n. fibers consistently show profiles of ER membranes and tubuli pertaining to the ER system. In some of these fibers the ER reaches a high degree of development. In Armadillidium there is a special type of n. fiber containing a regular sequence of transversally oriented cisternae. Arthropod fibers sometimes contain thin parallel filaments as well as typical ER elements.Mitochondria, small vesicles and dense granules are commonly found within the neuroplasm of the neuropilic fibers. It is important to note that in arthropods, microvesicles are not restricted to the terminal region of the nerve fibers but that they may also occur all along the fibers.Arthropod neurons are enveloped by a glial insulating capsule and therefore interneuron contacts may only occur at neuropile level. These contacts are of three different morphological types: cross contacts, longitudinal contacts and end-knob contacts. At the level of longitudinal and cross contacts the neuroplasm shows no increase in the number of microvesicles or mitochondria. In the end-knob contacts, on the contrary, large numbers of microvesicles appear concentrated in the pre-synaptic fiber only, and occasionally in both fibers the pre-synaptic and the post-synaptic.It is maintained that funcional interneuron connections may result not only from contacts between fibers containing vesicles, but also between fibers in which vesicles are absent.     

An immunofluorescence microscopical study of the neurofilament triplet proteins, vimentin and glial fibrillary acidic protein within the adult rat brain

A collection of antibodies specific to different intermediate filament proteins were applied to frozen sections of adult rat brains. The relative distribution of these proteins was then studied using double label immunofluorescence microscopy. Antibodies specific to each of the neurofilament "triplet" proteins (of approximate molecular weight 68 K, 145 K and 200 K) stained exclusively neuronal structures. The distribution of these three antigens was in general identical, except that certain neurofilament populations such as those in the dendrites and cell bodies of pyramidal cells of the hippocampus and cerebral cortex, contained relatively little if any 200 K protein. Some neurone populations, such as the granule cells of the cerebellar cortex, could not be visualized by neurofilament antibodies, indicating that neurofilaments may not be essential for function of all neurones in vitro. Antibodies to GFA and vimentin stained an entirely different population of processes, none of which stained with any of the neurofilament antibodies. Vimentin antibody stained sheath material around the brain, a monolayer of ependymal cell bodies lining the ventricles, fibrous material associated within the choroid plexus, the walls of blood vessels and capillaries, and the processes of cells in certain regions. GFA antibody stained a second layer of sheath material under the vimentin layer, and numerous processes visible throughout the brain. Some specific populations of GFA-positive processes proved to stain also with vimentin. These included the processes of Golgi "epithelial" cells (Bergmann glial fibres), those of certain astrocytes in bundles of myelinated fibers. In addition, some processes apparently derived from ependymal cells proved to stain for both vimentin and GFA, whilst other could only be reliably visualized by vimentin alone. These results are discussed in terms of the previously described morphological characteristics of the various cell types of the brain.     

Ultrastructural and immunohistological characterization of a cell culture model for the study of neuronal-glial interactions.

The differentiation of reaggregating cell cultures of dissociated cerebellar cells from 3- and 6-day-old mice was analyzed by electron microscopy of and immunofluorescence to the glial fibrillary acidic protein (GFA) at intervals between 8 hr and 20 days in vitro. Aggregates in culture for 8 hr were composed of 8–12 undifferentiated cells that were negative for the GFA protein and indistinguishable from each other by electron microscopy. Some cells with extended processes and a few immunofluorescent cells had already appeared after 24 hr in vitro, and the elaboration of a complex neural ultrastructure was observed during the subsequent days. After 10 days in vitro the interior of the aggregate was occupied principally by neurons, the majority of which were granule neurons, and regions of neuropil containing many synaptic complexes. Glial fibers with intense immunofluorescence to GFA were present throughout the aggregates but were mainly concentrated at the periphery. Large unidentified cells protruded from the surface. The subsequent days in culture evidenced a decline in the neuronal character of the aggregates with a concomitant increase in fibrous neuoglia. We suggest that factors controlling neuronal-glial interactions and fibrous gliosis are amenable to analysis in this tissue culture system.     

Enhanced neuronal expression in reaggregating cells of mouse cerebellum cultured in the presence of poly-L-lysine

Both neuronal and glial cell differentiation occur in reaggregating cell cultures of mouse cerebellums, as evidenced by electron microscopy and immunofluorescence to the glial fibrillary acidic protein (GFA). However, after the initial 10 days in culture a process occurs in which the neuronal cells degenerate while glial cells predominate. We have found that when poly-l-lysine is added to the culture medium either for the entire culture period or during the latter days of culture, i.e., Days 4 through 10, the neuronal character is stabilized, as evidenced by acetylcholin-esterase levels and electron microscopy, while the gliosis is inhibited. Culturing reaggregating cells in poly-l-lysine containing medium from Days 0 through 4 has no inhibitory influence on the gliosis observed on Day 10. Cerebellar cells cultured as monolayers on plastic surfaces coated with poly-l-lysine express an intense immunofluoresence with antisera to GFA as do cells grown on uncoated flasks. The data suggest that poly-l-lysine in reaggregating cell cultures stabilizes the neuronal cells by some unknown mechanism. It is postulated that a stable neuronal population reduces the trend toward gliosis in cerebellar aggregates.     

Neuronal and Glial Marker Proteins in the Evaluation of the Protective Action of MK 801

A quantitative dot immunobinding procedure was used to quantify glial [the S-100 protein and the glial fibrillary acidic (GFA) protein] and neuronal (the 68- and 200-kDa neurofilament polypeptides, neuron-specific enolase, and neuronal cell adhesion molecule) markers. A single intraperitoneal administration of 10 mg/kg of MK 801 blocked the increase of glial parameters and the decrease in content of neuronal marker proteins that occurred as the response to an N-methyl-D-aspartate (NMDA) lesion in the rat hippocampus. The degradation products of GFA protein and the 68-kDa neurofilament polypeptide that were induced by the NMDA lesion did not appear after MK 801 treatment. This study shows that brain-specific proteins are a set of precise tools for the evaluation of neuroprotective effects of antagonists to excitatory amino acids.     

Neuroprotection by α-Lipoic Acid in Streptozotocin-Induced Diabetes

Glial cells provide structural and metabolic support for neurons, and these cells become reactive to any insult to the central nervous system. The streptozotocin (STZ) rat model was used to study glial reactivity and the prevention of gliosis by alpha-lipoic acid (alpha-LA) administration. The expression of glial fibrillary acidic protein (GFAP), S100B protein, and neuron specific enolase (NSE) was determined as well as lipid peroxidation (LPO) and glutathione (GSH) levels in some brain tissues. Western blot analyses showed GFAP, S100B, and NSE levels significantly increased under STZ-induced diabetes in brain, and LPO level increased as well. Administration of alpha-LA reduced the expression both of glial and neuronal markers. In addition, alpha-LA significantly prevented the increase in LPO levels found in diabetic rats. GSH levels were increased by the administration of alpha-LA. This study suggests that alpha-LA prevents neural injury by inhibiting oxidative stress and suppressing reactive gliosis.     

An ultrastructural analysis of cell and matrix differentiation during early limb development in Xenopus laevis

Early development of the hind limb of Xenopus (stages 44–48) has been analyzed at the level of ultrastructure with emphasis on differentiation of extracellular matrix components and intercellular contacts. By stages 44–45, mesenchyme is separated from prospective bud epithelium by numerous adepidermal granules in a subepithelial compartment (the lamina lucida), a continuous basal lamina and several layers of collagen (the basement lamella). Tricomplex stabilization of amphoteric phospholipid demonstrates that each adepidermal granule consists of several membranelike layers (electron-lucent band 25–30 Å; electron-dense band 20–40 Å), which are usually parallel to the basal surface of adjacent epithelial cells. Collagen fibrils are interconnected by filaments (35 Å in diameter) which stain with ruthenium red. Epithelial cells possess junctional complexes at their superficial borders, numerous desmosomes at apposing cell membranes and hemidesmosomes at their basal surface. Mesenchymal cells predominantly exhibit close contacts (100–150 Å separation) with few focal tight junctions at various areas of their surface. By stages 47–48, adepidermal granules are absent beneath bud epithelium and layers of collagen in the basement lamella lose filamentous cross-linking elements. Filopodia of mesenchymal cells penetrate the disorganized matrix and abut the basal lamina. Hemidesmosomes disappear at the basal surface of the epidermis and mesenchymal cells immediately subjacent to epithelium exhibit focal tight junctions and gap junctions at their lateral borders. These structural changes may be instrumental in the epitheliomesenchymal interactions of early limb development. Degradation of oriented collagenous lamellae permits direct association of mesenchymal cell surfaces (filopodia) with surface-associated products of epithelial cells (organized into the basal lamina). Development of structural pathways for intercellular ion and metabolite transport in mesenchyme may coordinate events specific to limb morphogenesis.     

INCORPORATION OF 32P INTO THE PHOSPHOLIPIDS OF NEURONAL AND GLIAL CELL ENRICHED FRACTIONS ISOLATED FROM RABBIT CEREBRAL CORTEX: EFFECT OF NOREPINEPHRINE

Abstract— Glial cells isolated from rabbit cerebral cortex contained approximately one-third more phospholipids per unit protein than the neuronal cell bodies. The pattern of individual phospholipids was rather similar in both cell types. The incorporation of intracisternally administered ³²P into neuronal and glial phospholipid classes of rabbit brain was studied at intervals ranging from 5 to 60min. In general, for all investigated phospholipids the incorporation of the label was somewhat faster in neurons than in glial cells. Phosphatidylinositol showed the fastest and ethanolamine plasmalogen the slowest incorporation of ³²P in both neurons and glial cells. A lag phase of about 10 min could be observed before labelling of the glial phosphatidylcholine, phosphatidylethanolamine, ethanolamine plasmalogen, phosphatidylserine and sphingomyelin had occurred. Among the neuronal phospholipids a lag phase was found only for the labelling of the ethanolamine plasmalogen. Norepinephrine increased the incoropration of ³²P into phosphatidylinositol of both glia and neurons but had no effect on the specific radioactivity of ethanolamine plasmalogen and sphingomyelin. Labelling of phosphatidylcholine was slightly inhibited in both cell types by the administration of norepinephrine.     

Ultrastructural characteristics of anterior gut innervation of Gallus gallus

The enteric nervous system of the bird's anterior gut is very well developed. Myelin fibres are seen accompanying the nervous trunks up to the mucous layer. Glial cells duplicate the number of neurons in the myenteric plexuses. Their number decreases at the submucous plexuses, but it is always higher than the neurons. Isolated neurons are widely spread in the circular muscle coat accompanying the nervous trunks which can be inter and intrafascicularly located. Direct synaptic contacts with the soma neuronal membranes are very often seen. We have never observed synaptic specializations. The most prominent varicosities either in the peripheric nervous trunk axons or directly laying on the soma membranes are those containing peptidergic or mixed vesicles of cholinergic and peptidergic types. The neurons show big nuclei of different size and shape. Neighbouring smooth muscle cells show abundant caveolae near the nervous elements. Although we have not observed close contacts with glands, thin axon bundles spread near the glandular cells of the mucous layer.     

Sympathetic glial cells and macrophages develop different responses to Trypanosoma cruzi infection or lipopolysaccharide stimulation

Nitric oxide (NO) participates in neuronal lesions in the digestive form of Chagas disease and the proximity of parasitised glial cells and neurons in damaged myenteric ganglia is a frequent finding. Glial cells have crucial roles in many neuropathological situations and are potential sources of NO. Here, we investigate peripheral glial cell response to Trypanosoma cruzi infection to clarify the role of these cells in the neuronal lesion pathogenesis of Chagas disease. We used primary glial cell cultures from superior cervical ganglion to investigate cell activation and NO production after T. cruzi infection or lipopolysaccharide (LPS) exposure in comparison to peritoneal macrophages. T. cruzi infection was greater in glial cells, despite similar levels of NO production in both cell types. Glial cells responded similarly to T. cruzi and LPS, but were less responsive to LPS than macrophages were. Our observations contribute to the understanding of Chagas disease pathogenesis, as based on the high susceptibility of autonomic glial cells to T. cruzi infection with subsequent NO production. Moreover, our findings will facilitate future research into the immune responses and activation mechanisms of peripheral glial cells, which are important for understanding the paradoxical responses of this cell type in neuronal lesions and neuroprotection.     

Glial cells of the crayfish and their relationships with neurons. An ultrastructural study

1. Glial cells of the crayfish abdominal ganglia have been studied by transmission electron microscopy. Special attention is paid to the interrelationships between neurons and glial cells. Covers and hemocyte-related elements have also been considered. 2. Glial cells are identified by a common ultrastructure and close relationships with neurons. Four glial classes are considered, depending on their morphology, the compartment of neurons they ensheathe and neuron-glia interface. 3. Four ultrastructural classes of neurons are proposed. They differ in geometry and ultrastructure, as well as in glial covers (complexity and evaginations into the neuron somata). The morphology and organization of glial covers is specific for the neuron type they ensheathe. Specific glial covers do not differ in glia-glia communicatory structures. 4. The morphological and metabolical compartments of neurons are separated from the extracellular matrix or blood by specific glial systems. A system of two cells is interposed between neuron somata and hemolymph or the extracellular matrix. 5. Glial processes are crossed by membraneous tubular systems, at neuron perikarya and axons. Frequent gap junctions of varying area, density and number of IMP are found in the covers of neuron somata. 6. Neuron-glia interface bears numerous communicatory structures for both ionic and macromolecular exchange. They include junctions and transient modifications of membranes. Some of them suggest active transport mechanisms. 7. Modified endocytotic mechanisms seem to be responsible for the glia-to-neuron transfer of macromolecules as well as for the neuron-to-glia transfer of lamellar bodies. 8. The neuropil is divided into glomeruli (electrical or chemical) by glial processes and the trabeculae of the extracellular dense matrix. Neuron-glia membrane appositions have been found in electrical glomeruli. In chemical glomeruli, dense cored vesicles can release their content at neuron-neuron or neuron-glia intercellular cleft, at non-synaptic loci. 9. Neurons of type II contain peripheral complex Golgi systems, associated to subsurface cisternae and neuron-glia gap junctions, suggesting a cooperation of glial cells in specific macromolecular synthesis.     

Glial patterning during postembryonic development of central neuropiles in the brain of the honeybee

 Glial cells are involved in several functions during the development of the nervous system. To understand potential glial contributions to neuropile formation, we examined the cellular pattern of glia during the development of the mushroom body, antennal lobe and central complex in the brain of the honeybee. Using an antibody against the glial-specific repo-protein of Drosophila, the location of the glial somata was detected in the larval and pupal brain of the bee. In the early larva, a continuous layer of glial cell bodies defines the boundaries of all growing neuropiles. Initially, the neuropiles develop in the absence of any intrinsic glial somata. In a secondary process, glial cells migrate into defined locations in the neuropiles. The corresponding increase in the number of neuropile-associated glial cells is most likely due to massive immigrations of glial cells from the cell body rind using neuronal fibres as guidance cues. The combined data from the three brain regions suggest that glial cells can prepattern the neuropilar boundaries. Received: 3 November 1996 / Accepted: 7 February 1997     

Morphological characterization of a human glioma cell l ine

A human malignant continuous cell line, named NG97, was recently established in our laboratory. This cell line has been serially subcultured over 100 times in standard culture media presenting no sign of cell senescence. The NG97 cell line has a doubling time of about 24 h. Immunocytochemical analysis of glial markers demonstrated that cells are positive for glial fibrillary acidic protein (GFAP) and S-100 protein, and negative for vimentin. Under phase-contrast microscope, cultures of NG97 showed cells with variable morphological features, such as small rounded cells, fusiform cells (fibroblastic-like cells), and dendritic-like cells. However, at confluence just small rounded and fusiform cells can be observed. At scanning electron microscopy (SEM) small rounded cells showed heterogeneous microextentions, including blebs and filopodia. Dendritic-like cells were flat and presented extensive prolongations, making several contacts with small rounded cells, while fusiform cells presented their surfaces dominated by microvilli.