Cellular Interdigitation Research Articles

O-233 Cell-cell interactions within germ cell nests reveal the mechanism of primordial follicle formation in humans

Abstract Study question How interactions between human fetal female germ cells (hFGCs) and pre-granulosa cells (pre-GCs) affect the hFGCs transition from ovarian cord to primordial follicles (PFs). Summary answer Progressive engulfment of hFGCs by pre-GCs and establishment of cell-cell junctions and cell-cell adhesions between hFGCs and pre-GCs leads to PF formation in humans. What is known already In mice, development of FGCs is synchronized due to formation of cysts. Cysts comprise of sister FGCs that remain connected via intercellular bridges until germ cell nest breakdown is initiated and PFs are formed. In contrast, hFGCs differentiate asynchronously. In second trimester human ovaries, mitotic, RA-responsive, meiotic and oogonia hFGCs are present in designated parts of the ovary that corresponds to their differentiation stage. Meiotic hFGCs enter prophase I that is divided into 4 substages: leptotene, zygotene, pachytene, diplotene/dictyate. hFGCs in diplotene are ready to form a PF with pre-GCs and become dormant. Study design, size, duration We collected in total 17 second trimester human fetal ovaries between 14 and 22 weeks of gestation from elective abortions. We have analyzed cellular structures of hFGC and pre-GCs by transmission electron microscopy and investigated different cell adhesion and cell junction proteins by immunostaining. Participants/materials, setting, methods Materials: human fetal ovaries between 14 and 22 weeks of gestation. Methods: characterization of tissue morphology, immunofluorescence staining, whole mount staining, confocal microscopy, super-resolution microscopy, transmission electron microscopy (TEM), cell counting Main results and the role of chance Meiotic hFGCs formed germ cell cysts consisting of multiple sister hFGCs connected by intercellular bridges. Furthermore, we discovered that a subset of hFGCs formed syncytia containing several nuclei (2–4 nuclei). Syncytium formation was caused by cytokinesis failure when an in intercellular bridge failed to be formed. In addition, in some cases hFGC syncytia were connected to other syncytia or mono-nucleated hFGCs by an intercellular bridges. Those multinucleated syncytia did not display signs of apoptosis or morphological abnormalities. Investigation of physical interactions between hFGCs and pre-GCs revealed that as hFGCs enter prophase I, pre-GCs (FOXL2+, VIM+, CDH1+) form membrane protrusions (MPs) and progressively engulf them. First, CDH1+ and VIM+ domains within a pre-GC are mixed, but as engulfment progresses two distinct domains are formed. MP formation is possible due to mesenchymal-epithelial hybrid state and active Hippo pathway in pre-GCs. Inside the cords, hFGCs highly express CDH2, while CDH2 is lowly expressed in pre-GCs. At diplotene, hFGCs (TP63+) downregulate CDH2 and upregulate CDH1. Expression of CDH1 in both diplotene hFGC and pre-GCs causes increase of cell-cell adhesion and enables PF formation. In addition, progressive formation of tight junctions (TJP1+) and cellular interdigitations between pre-GCs and meiotic hFGCs reinforce PF structure. Limitations, reasons for caution Due to technical limitations and lack of in vitro culture protocols that support meiotic hFGCs development and survival as well as human PF formation in vitro, we cannot experimentally validate roles of cadherins and other cell junction and cell adhesion proteins in PF formation Wider implications of the findings In depth understanding of PF formation process in terms of physical interactions between hFGCs and pre-GCs will be crucial to design in vitro conditions that will support PF formation and our findings will be key to achieve assembling of PFs in artificial human oocytes (gametes) created from pluripotent stem cells. Trial registration number not applicable

Crop Morpho-Histological Peculiarities in Domesticated Pigeons (Columba livia domestica), Cattle Egret (Bubulcus ibis) and Domesticated Ducks (Anas platyrhynchos domestica)

The crop architecture varies among different avian species consistent with their feeding habits. Therefore, twenty-one of mature healthy male pigeons (granivorous), cattle egrets (carnivorous), and ducks (omnivorous), seven per species, were utilized for the histological, immunohistochemical, and ultrastructural analyses. Histologically, the mucosal folds were covered by non-keratinized stratified squamous epithelium in the three investigated species. The mucosal glands are only peculiar to the crops of the cattle egrets as well as the ducks, while the pigeons' crops were devoid of any secretory units. Morphometrically, the optical densities of the Alcian blue (AB) and periodic acid Schiff (PAS) reactions in the secretory glands, the area % of collagen fibers, the thickness of the tunica musculosa were measured and declared significant differences among the three studied avian species. Immunohistochemical reaction revealed ki-67 immuno-positive reactivity in the nuclei of basal cell layers of the crop epithelium in pigeons only. Regarding ultrastructure investigation, the covering epithelium of all studied species had been shown to be the basal layer of cuboidal to tall columnar cells with desmosomal junctions at the level of their cellular interdigitation, intermediate layers of large irregular polygonal cells with obviously increased cytokeratin filaments especially in the ducks and cellular interdigitation in between. The superficial layer of flat-shaped cells in the three investigated species manifested by desmosomal junctions in between but at the level of the surface squamous cells of the superficial layer, a fine lateral process is only inspected in the pigeons. In conclusion, the crop glands characterized by supranuclear electron dense secretory granules in the cattle egrets and electron-lucent in ducks.

Cytological and Ultrastructural Findings of Mucinous Tubular and Spindle Cell Carcinoma: A Case Report

The 2016 World Health Organization Classification of Tumors recognized mucinous tubular and spindle cell carcinoma (MTSCC) as a rare histological variant of renal cell carcinoma (RCC). To date, a limited number of studies have reported this tumor concerning histology, and only a handful of those have reported cytological and ultrastructural features. This study aims to report a case of MTSCC in a female patient in her early 50s who presented with a chief complaint of gross hematuria, and to discuss relevant cytological and ultrastructural findings. Imaging revealed a 30 mm tumor in the left kidney, suggesting the presence of RCC, and left nephrectomy was performed. In addition, scratched cytology of tumor surface revealed clusters of tumor cells with clear cell boundaries extending two-dimensionally in papillary or dendritic arrangements with interspersed blood vessels and stromal components that were stained light green. Histologically, the tumor was composed of small and uniform cells arranged in elongated tubular architectures and spindle fascicles with occasional papillary structures; the stroma was acidophilic and contained colloidal substances that tested positive for Alcian blue and periodic acid-Schiff stain. Concordant with immunohistochemical results, the tumor was diagnosed with MTSCC. Furthermore, transmission electron microscopy revealed tubular structures with microvilli, abundance of mitochondria in the cytoplasm, cellular interdigitations and widened interstitium.

Auxin Binding Protein 1 (ABP1): a matter of fact.

Science is a global community enterprise.Collectively,we contribute to building the knowledge tower by using bricks and mortar that should be strong,solid,and long lasting.A brick with defects may lead this tower,or a part there of,to collapse.When that happens,the community suffers,laying waste countless dedicated hours of work,especially by students and postdoctoral scholars.This is the reason ary

Myxoid meningioma: a rare metaplastic meningioma variant in a patient presenting with intratumoral hemorrhage

Myxoid (metaplastic) meningioma is a rare WHO Grade I meningioma subtype arising from the leptomeninges. It has unique Alcian blue stromal staining and distinctive cellular interdigitations, junctional complexes, and nucleolar pseudoinclusions on ultrastructural pathology that help to distinguish it from other meningioma variants. The authors describe the case of a rare left middle fossa, extraaxial myxoid meningioma in a 50-year-old woman to emphasize the important histological characteristics and observations essential for making a precise diagnosis. To their knowledge this is the seventh reported case of a myxoid meningioma in the literature and the sixth case in an adult; however, it is the first reported instance of myxoid meningioma in a patient presenting with intratumoral hemorrhage.

Mucous Cells in the Gills of Snow Trout, Schizothorax curvifrons Heckel (Pisces, Cypriniformes, Cyprinidae)

The ultrastructural and histochemical characteristics of mucous cells and their functional significance in the gills of Schizothorax curvifrons were studied with the objectives that this study could serve as a basis for further studies in relation to histopathology, aquatic toxicology and stress conditions in aquaculture and natural or polluted environments. The electron microscopical analysis of the gills following fixation in Karnovsky's fluid and post fixation in osmium tetraoxide revealed that the mucous cells were covered by epithelial cells and their major part was occupied by firmly arranged large mucous globules of different electrodensities, displacing their nuclei towards the basal position. The cytoplasmic extensions and islets among the mucous globules and the cellular interdigitations between mucous cells and neighbouring cells were present and the release of mucin by exocytosis was also observed. Utilizing histochemical techniques that included methods of periodic acid Schiff (PAS) reaction and alcian blue method (AB pH 2.5) for localization of neutral and acid mucopolysaccharide secreting mucous cells showed that the gill epithelium was lined by mucous cells which were found to be PAS and AB positive, confirming the presence of both neutral mucous cells secreting hexose containing mucosubstances and acidic mucous cells secreting weakly acidic sulphated mucosubstances and sialomucins. The possible roles of the mucous cells and the products secreted by them have been discussed in this paper.

Cell Surface- and Rho GTPase-Based Auxin Signaling Controls Cellular Interdigitation in Arabidopsis

Auxin is a multifunctional hormone essential for plant development and pattern formation. A nuclear auxin-signaling system controlling auxin-induced gene expression is well established, but cytoplasmic auxin signaling, as in its coordination of cell polarization, is unexplored. We found a cytoplasmic auxin-signaling mechanism that modulates the interdigitated growth of Arabidopsis leaf epidermal pavement cells (PCs), which develop interdigitated lobes and indentations to form a puzzle-piece shape in a two-dimensional plane. PC interdigitation is compromised in leaves deficient in either auxin biosynthesis or its export mediated by PINFORMED 1 localized at the lobe tip. Auxin coordinately activates two Rho GTPases, ROP2 and ROP6, which promote the formation of complementary lobes and indentations, respectively. Activation of these ROPs by auxin occurs within 30 s and depends on AUXIN-BINDING PROTEIN 1. These findings reveal Rho GTPase-based auxin-signaling mechanisms, which modulate the spatial coordination of cell expansion across a field of cells.

Cell Surface- and Rho GTPase-Based Auxin Signaling Controls Cellular Interdigitation in Arabidopsis

Cell Surface- and Rho GTPase-Based Auxin Signaling Controls Cellular Interdigitation in Arabidopsis

Pathogenesis of some neurological immune ultrastructural and morphometrical observations on rat thymus

Numerous studies on neuro-immuno-modulation indicate that the thymus is involved in many neurological diseases, including experimental allergic encephalomyelitis (EAE). Twenty Lewis rats were induced for EAE. At X, XII, XX and XXX days post-inoculation the animals were killed, and the thymus was recovered and harvested. Specimens of thymus were submitted to morphological light microscopy analysis (1% toluidine blue) and ultra-structural analysis (transmission electron microscopy). Significant morphometric data were collected by examining the images quantitatively and by statistically analysing the values. Our results show that the microenvironment of the thymus is severally involved in acute EAE. Thymocytes and reticular epithelial cells show many changes which are closely related to the pathogenesis of EAE. In particular we observed: (1) inside the cell an increase in intra-cytoplasmic vacuoles, and changes in the thickness of the nuclear membrane, mitochondria, rough endoplasmic reticulum, cellular inter-digitations and cellular electron-density; (2) outside the cell an increase in pericellular translucent halo, intercellular spaces, intercellular contacts and apoptotic and necrotic figures. The evidence of a thymic role in MS may suggest the intriguing therapeutic concept of thymectomy in the management of this neurological disease.

ENCAPSULATION OF MYXOBOLUS PENDULA (MYXOSPORIDIA) BY EPITHELIOID CELLS OF ITS CYPRINID HOST SEMOTILUS ATROMACULATUS

Spores of Myxobolus pendula develop within the cores of complex cysts on the gill arch of creek chub Semotilus atromaculatus. Adjacent to, and surrounding, the spores are concentric layers of stratified interdigitating cells, whose nature was examined by transmission electron microscopy and by immunohistochemical and molecular biological techniques. In situ hybridization data using parasite-derived ribosomal DNA as a probe indicate that infection leads to the encapsulation of developing plasmodia by host immune cells that form an epithelioid granuloma. Epithelioid cell-cell adhesion is effected by desmosomes anchored intracellularly to cytokeratin intermediate filaments. High levels of alkaline phosphatase activity are associated with regions of cellular interdigitation. The granuloma may serve to limit the spread of the parasite to surrounding tissues but does not appear to inhibit diffusion of oxygen and nutrients to the developing spores.

The role of epithelial remodelling in tooth eruption in larval zebrafish.

Based on light and transmission electron-microscopic observations on erupting first-generation teeth in the zebrafish, Danio rerio, we propose a biphasic mechanism for tooth eruption: (1). formation of an epithelial crypt prior to eruption of the tooth, possibly as a result of constraints in the epithelium resulting from the growth of adjacent tooth germs, and (2). detachment of cellular interdigitations both within the pharyngeal epithelium, at the pharyngeal epithelium/enamel organ boundary, and between the outer and inner dental epithelium, resulting in the exposure of the tooth tip in the crypt, immediately after tooth ankylosis. Later, further detachment of interdigitations between the inner and the outer enamel epithelium unfolds the epithelium even more and leads to a more pronounced exposure of the tooth tip. The presence of small patches of non-collagenous matrix on the outer surface of the tooth close to where it merges with the attachment bone is interpreted as a device to prevent complete detachment of the enamel organ. The biphasic nature of the mechanism for tooth eruption is supported by observations on in vitro cultured heads. First-generation teeth develop normally and crypts are formed, as under in vivo conditions, but the teeth fail to erupt. Taken together, our observations suggest that epithelial remodelling plays a crucial role in eruption of the teeth in this model organism.

Histochemical and ultrastructural features of the lingual epithelium of the rat snake (Elaphe climacophora)

The histological, ultrastructural and histochemical characteristics of the lingual epithelium of the rat snake (Elaphe climacophora) were investigated by light and transmission electron microscopy. The cells in the beta-layer of the epithelium of the bifurcated apex were filled with beta-keratin fibers and an amorphous matrix. Round projections covering the surface of the epithelial cells, namely, microfacets which contained pale granules, were clearly visible on the outer faces of Oberhäutchen cells on the epithelium, and they were identified as fine granules filled with lipid. These granules might play an important role as a coating on the surface of the bifurcated lingual apex. The lipid on the surface of the lingual apex might also serve to trap and retain odorant molecules. Keratohyalin-like granules were distributed within the a-layer of the epithelium of the bifurcated apex of the tongue in the resting phase and cellular interdigitation was well developed in this region. Evidence of a shedding line was apparent under the light microscope in the cleft between outer and inner epithelial generations. The epithelial surface of the body of the tongue appeared suitable for retention of odorant and other molecules.

Reexamination of the Fine Structure of the Pancreatic Duct System in the Rat: a Proposal of Morphological Segmentation

The pancreatic duct system is generally regarded as a uniform conduit lined by principal cells that secrete bicarbonate-rich fluid. However, previous studies have suggested the morphological and functional heterogeneity of these cells. We reexamined the fine structure of the principal cells in the rat pancreas by both scanning electron microscopy after exposure of the cells with NaOH maceration, and by transmission electron microscopy according to a conventional method. The specimen was also observed by immunohisochemistry with a monoclonal antibody OS-3 recognizing the renal type B intercalated cell. The ducts in the rat pancreas were lined by a simple epithelium devoid of basal cells, and arborized dichotomously. The duct system consisted of two parts: a fine proximal duct and thicker distal duct. Principal cells in the latter, as compared with those in the former, displayed relatively numerous microplicae on the basolateral membrane and abundant mitochondria in the cytoplasm. The distal duct cells were also unique in their possessing secretory canaliculi opening into the main lumen as well as thick lateral processes for cellular interdigitation. These findings suggest that the distal duct has a greater ability to transport ions than does the proximal one. The antibody OS-3 preferentially labeled the distal duct cells, augmenting their distinction from the proximal duct cells.

Altered expression of type II sodium/phosphate cotransporter in polycystic kidney disease.

Renal phosphate (Pi) absorption is mediated via the type II sodium/Pi cotransporter (NaPi-2) in the brush border membrane (BBM) of proximal tubules. Simultaneous detection of NaPi-2 mRNA by in situ hybridization and of NaPi-2 immunoreactivity by immunohistochemistry was performed to investigate the distribution of the cotransporter in healthy control rats and during progression of autosomal dominant polycystic kidney disease (ADPKD). The purpose of the study was to disclose a relation between proximal tubular cell differentiation and NaPi-2 expression. In controls, NaPi-2 expression was present in the entire proximal tubule. In the Han:SPRD (cy/+) model for ADPKD, the proximal nephron is primarily affected by the cystic changes. Epithelial proliferation and impaired epithelial-matrix interaction result in a loss of cell differentiation that eventually leads to cystic enlargement of the nephron. Normal expression of NaPi-2 in this model was found only in tubules with intact BBM. Loss of BBM and cellular interdigitation were paralleled by the loss of NaPi-2 in situ hybridization and immunoreactive signals. These changes were moderate and focal in 2-mo-old rats and generalized all over the cortex after 8 mo. Advanced renal damage in the older PKD group was associated with mild phosphaturia, which suggests functional insufficiency of tubular NaPi-2 reabsorption. These data show how proliferative changes and loss of tubular epithelial differentiation in ADPKD may prevent functional expression of the NaPi-2 system in the proximal tubule in a rapidly progressive manner. NaPi-2 in proximal tubule BBM is suggested to play an important role in impaired tubular absorption of Pi in renal disease.

In situ localization of anion exchanger-2 in the human kidney

Na+-independent anion exchangers (AE) are a family of membrane carriers that mediate the electroneutral exchange of Cl- for HCO3- ions across plasma membranes. They are involved in intracellular pH and cell volume regulation as well as in transepithelial acid-base transport. While anion exchanger-1 (AE1) has been localized previously in the human kidney, thus far there has been no definite report on anion exchanger-2 (AE2) in this human tissue. Accordingly, immunohistochemistry was carried out on surgical specimens of the human kidney (fixed in formalin and embedded in paraffin), using a specific AE2 monoclonal antibody. Strong immunostaining was observed at the basolateral membrane of cells of thick ascending limbs and distal convoluted tubules, colocalizing with the basal membranous labyrinth of cellular interdigitations, typical of these segments. In fact, AE2 staining was attenuated at the macula densa, where basal infoldings are scarce. Additionally, in situ hybridization experiments on formalin-fixed tissue demonstrated the presence of AE2 mRNA in the same segments of the distal nephron. On the other hand, control immunohistochemistry with a monoclonal antibody against AE1 gave the expected immunoreactivity at the basal pole of the type A intercalated cells of connecting tubules and cortical collecting ducts, and in erythrocytes. Our results indicate that, depending on the nephron segment and corresponding cell types, AE1 and AE2 proteins are differentially involved in the Na+-independent exchange of Cl- for HCO3- at the basolateral membrane of polarized kidney epithelial cells.

Three-dimensional microanatomy of perineuronal proteoglycan nets enveloping motor neurons in the rat spinal cord.

Spinal motor neurons possess reticular coats of extracellular matrix proteoglycans on their somata and proximal dendrites. In order to define the anatomical background of the network, spatial relationships of the perineuronal proteoglycans with synaptic boutons and astrocyte processes were analyzed in rat motor neurons by TEM after histochemical detection of the substances with cationic iron colloid, and by SEM after exposure of the cytoarchitecture with NaOH maceration. Narrow intercellular channels filled with proteoglycan were found to extend along the surface of the neurons to form a homogeneous network of a mesh size of about 1 microm. The system of perineuronal channels consisted of two parts: a primary intervaricose net which meandered among synaptic boutons on the surface of the motor neuron, and secondary subvaricose nets which irrigated interfaces between larger boutons and the neuron. No elements in the perineuronal cytoarchitecture coincided with the meshwork of proteoglycan, indicating the involvement of postsynaptic factors in the distribution of the substance. Thin astrocyte processes surrounding the neurons formed a distinct network with heterogeneous meshes corresponding to boutons of various sizes. The perineuronal glial nets extended their surface area in contact with the intervaricose nets of proteoglycan by complex cellular interdigitations. The subvaricose nets of proteoglycan compartmentalized multiple synapses on large boutons, suggesting an involvement in the division of the synapses during development.

An ultrastructural study of the dorsal lingual epithelium of the rat snake, Elaphe quadrivirgata

The histological characteristics and ultrastructure of the dorsal lingual epithelium of the rat snake, Elaphe quadrivirgata, were investigated by light microscopy and scanning and transmission electron microscopy. Most of the surface of the bifurcated part of the tongue was relatively smooth. Dome-shaped, hemispherical bulges were compactly arranged on the epithelial cell surface of the basal area of this region. Intercellular borders were clearly recognizable as striations. Microridges were densely distributed on the epithelial cell surface of the lingual body. Intercellular borders were thickened. A keratinized layer was clearly visible in the epithelium of the anterior bifurcated area, namely, at the apex of the tongue. Although keratohyalin granules were not found in any layer of the epithelium in this area, the cells of the surface layer were filled with keratin filaments. The dorsal lingual epithelium of the posterior area, namely, the lingual body, did not show any evidence of keratinization. Each cell on the surface side still had a large, oval nucleus and intact organelles, such as mitochondria, rough endoplasmic reticulum, ribosomes, tonofibrils, and tonofilaments. Cellular interdigitation was evident between adjacent cells and clear microridges or microvilli were observed on the cell membranes on the free-surface side of cells located in the surface layer. The phylogenetic relevance of these findings is discussed.

Hemispheric-apex cells in the mantle epithelium ofLoligo vulgarisandLoligo forbesihatchlings and juveniles: a transmission electron microscope study

Summary Hemispheric-apex cells (HC), which are situated at the intersection of the three branches of Hoyle's organ in Loligo vulgaris and Loligo forbesi embryos, were investigated using light- and transmission-electron microscopy. The surface membrane of the HC exhibits many microvilli. Desmosomes between HC were distributed in patches along the plasma membrane. These desmosomes were also found between HC and the hatching enzyme producing cells of Hoyle's organ (CHO) and between neighbouring CHO. The area below the most apical desmosomes shows many cellular interdigitations. One (or several) small Golgi apparatus and many mitochondria were found in the cell centre. Mitochondria were also abundant in the most apical part of the cell. In the HC the nucleus is situated near the cell centre. The cell shape of the central part of the cell usually corresponds grossly to the nuclear form. Endoplasmic reticulum was found near the nucleus and in the lowermost part of the cell, but is never as abundant as in the CH...

The Process of Secretion in Swine Apocrine Sweat Glands

The secretory process in swine apocrine sweat glands were studied by electron microscope. The sweat appeared to result from three different mechanisms: (a) fluid transport, probably involving a region of complex cellular interdigitations adjacent to the basement membrane, (b) exocytosis of vesicles, which in this species seem to be derived from the Golgi apparatus and finally (c) apocrine secretion.

Lymphomatoid Granulomatosis in a Beagle Dog

Lymphomatoid granulomatosis is a rare human disease that has been recently reported in young dog^.',^.^.^ Lymphomatoid granulomatosis, as originally described in human beings by Liebow et al.,5 is an angiocentric and angiodestructive lymphoreticular proliferative and granulomatous disease that predominantly involves the lungs. Pathologically, it is a peculiar angiocentric lymphoid lesion with a lymphomalike appearance (lymphomatoid) in some areas and necrosis (granulomatosis) in others. The proliferative infiltrate is composed of atypical lymphoreticular cells admixed with variable proportions of lymphocytes, plasma cells, and histiocytes. A clinically normal, 8-month-old, male Beagle was not included in drug safety studies owing to a persistent eosinophilia (3.2-4.2 103/mm3). The dog was euthanatized at 15 months of age as a result of a sudden deterioration of its condition: a marked abdominal enlargement, a 3.7 kg increase in body weight during the last 2 weeks, and severe respiratory problems, including coughing. At necropsy, the abdomen contained about 2.5 liters of a clear fluid; the liver was congested. The accessory lobe of the right lung was enlarged (1 2 x 13 cm), firm, and pale yellow. The overlying pleura was smooth and dull. Cut sections revealed coalescing multinodular pale tissue centered around bronchi that compressed the remaining lung peripherally. Grey or dark-red patches were scattered throughout all lobes. A white, firm tissue replaced the markedly enlarged tracheobronchial lymph nodes (5 x 3 cm) and was found focally in the sternal lymph node. Other organs were unremarkable. Tissues were fixed in 10% neutral buffered formalin, routinely processed to paraffin sections, and stained with hematoxylin and eosin, and Gomori's trichrome stain with Weigert's stain for elastin fibers. Sections were stained immunocytochemically by the peroxidase anti-peroxidase technique using rabbit anti-bovine S100 protein (Dakopatts, Issy-les-Moulineaux, France) with a silver amplification (Amersham, Les Ulis, France), and by peroxidase-labeled protein A (New England Nuclear, Paris, France) with trypsinization, using goat anti-human a1 -anti-trypsin, goat antihuman lysosyme followed by a rabbit anti-goat immunoglobulin G (Nordic, Le Perray en Yvelines, France). For negative controls, duplicate sections were incubated with normal, nonimmune rabbit or goat serum instead of the primary antibody. Lymph node and brain were used as positive controls. Formalin-fixed portions of the accessory lobe of the right lung were divided into 1 mm3, immersed in 2.5% glutaraldehyde in 0.1 M phosphate buffer (pH 7.37), post-fixed in 2% osmium tetroxide, and embedded in epoxy resin. U1trathin sections were stained with uranyl acetate and lead citrate and examined with a Zeiss EM109 electron microscope. Microscopically, the tumor was composed of sheets of dense infiltrates of atypical pleomorphic lymphoid cells that obliterated normal structures (Fig. 1). Most cells were polygonal to oval and had a faintly basophilic, moderately abundant cytoplasm. Nuclei varied from rounded to slightly elongated. Chromatin was predominantly stippled and occasionally coarse. Many cells had up to four round amphophilic nucleoli. There were atypical vesicular or cleft nuclei. Scattered binucleated cells were present. The fine fibrovascular stroma was predominantly infiltrated by eosinophils, fewer plasmocytes, and occasional Mott cells. Individual cell necrosis was observed throughout the tissue. No immunoreactivity for lysozyme, a-I-antitrypsin or S-100 protein was seen in tumor cells. Many large blood vessels were infiltrated by pleomorphic lymphoid cells and eosinophils. Their centers were occupied by a variously-shaped fibrous tissue that delineated vascular channels and was connected to the fibrous wall that merged with the surrounding stroma (Fig. 2). They had no elastic lamina. There were occasional epithelial remnants of airways and necrotic foci up to 3 mm in diameter. The overlying pleura was thickened and fibrotic. Sections at the margins of the tumor revealed that the process originated from pulmonary arteries and compressed adjacent airways (Fig. 3). The intimae of some affected pulmonary arteries were thickened by edema, fibrosis, eosinophils, and atypical lymphoid cells. Histologic lesions of allergic bronchitis3 with a few, small, foreign-body granulomas corresponded to the patches seen at necropsy in the other lobes. Blood vessels in these areas were spared. The sternal lymph node was multifocally invaded by tumor cells. Histologic liver changes were consistent with acute passive congestion. Heart, kidney, spleen, thyroid, and muscle were unremarkable. Ultrastructurally, tumor cells lacked intercellular junctional complexes and pericellular basement membrane material (Fig. 4). A limited amount of cellular interdigitations was observed. Nuclei were ovoid or variably folded and had large, often multiple, nucleoli. The cytoplasm contained moderately abundant profiles of granular endoplasmic retic-