Search

The role of protein kinase C (PKC) in the regulation of the cytoskeleton of epithelial cells with tightly sealed contacts, poor contacts, and without contacts were investigated by incubating them with a protein kinase C activator phorbol myristoyl acetate (PMA). The morphology and organization of the membrane skeleton and stress fibers as well as the localization of an actin-bundling PKC substrate MARCKS in confluent MDCK cells originating from the distal tubulus of dog kidney, LLC-PK1 cells originating from the proximal tubulus of pig kidney, src-transformed MDCK cells, epidermoid carcinoma A431 cells, and MDCK cells grown in low calcium medium (LC medium) in low density were visualized with phase contrast and immunofluorescence microscopy. Four different responses to the PMA-treatment in actin-based structures of cultured epithelial cells were observed: 1) disintegration of the membrane skeleton in confluent MDCK cells; 2) depolymerization of the stress fibers in confluent MDCK and LLC-PK1 cells; 3) formation of the membrane skeleton in A431 cells, and 4) formation of the stress fibers and membrane skeleton in LC-MDCK cells. Thus, it seems that in fully confluent tightly sealed epithelium, activation of PKC has a deleterious effect on actin-based structures, whereas in cells without contacts or loose contacts, activation of PKC by PMA results in improvement of actin-based cytoskeletal structures. The main difference between the two kidney cell lines used is their selectivity to ion transport: the monolayer of LLC-PK1 cells is anion selective and MDCK cells cation selective. We propose a model where alterations in the ionic milieu within the MDCK cells by means of cation channels affect the disintegration of the membrane skeleton. The distribution of MARCKS followed the distribution of fodrin in both cell lines upon PMA-treatment, suggesting that phosphorylation of MARCKS by PKC may contribute in the regulation of the integrity of the membrane skeleton., (Copyright 1999 Wiley-Liss, Inc.)

The signaling pathways from an activation of protein kinase C (PKC) by phorbol myristate acetate (PMA) to the rearrangement of actin-based cytoskeleton and membrane skeleton of epithelial MDCK cells were studied by visualizing the cytoskeletal organization with immunofluorescence microscopy and by measuring intracellular pH, sodium ion concentration and membrane potential with the aid of fluorescent intracellular indicators. Upon PMA treatment the MDCK cells lost their cubic shape and acquired a spindle-like morphology. The stress fibers were depolymerized, and fodrin, the main component of the membrane skeleton, was released from the lateral walls to the cytosol. These changes were accompanied by depolarization of the cells, decrease in the intracellular pH and sodium ion concentration. In order to test the mutual correlation between the PMA-induced alterations we treated the cells with PMA in the presence of channel inhibitors or ionophores and in defined media. The effects of PMA on the membrane skeleton and morphology could be reversed in media lacking Na+ or K+ ions or by hyperpolarizing agents, dimethylamiloride and valinomycin, suggesting that the effects of PMA on the cytoskeleton were dependent on the ion gradients and membrane potential across the cell membrane. Moreover, the morphological changes and instabilization of the membrane skeleton of MDCK cells took place spontaneously without PMA in depolarizing conditions, in potassium gluconate buffer. We suggest that the membrane potential across the cell membrane of MDCK cells together with the activity of amiloride-sensitive cation transporters transmits signals in the protein kinase C (PKC) pathway leading from activation of PKC to fibroblast-like morphology and cytoplasmic localization of membrane skeleton components, features characteristic for cancer cells.

The commonly accepted markers for amyliod are a fibrillar ultrastructure and congophilia combined with green birefringence when viewed under polarized light. Although a number of cells have been implicated in amyloid formation the detailed events leading to accumulation of amyloid are still unknown. Previous studies have suggested that amyloid shares antigenic properties with cytoskeletal intermediate (10 nm) filaments and connective tissue microfibrils. In the present study we show that such filaments present in fibroblasts and lymphoid cells have an affinity for Congo red, exhibit the typical green birefringence and are ultrastructurally indistinguishable from amyloid fibrils. Intermediate filaments of cultured fibroblasts depleted of nutrient medium retained the Congo red birefringent property and typical ultrastructure despite cell damage. Our observations suggest that amyloid deposits may form locally through abnormal accumulation of bundles of cytoskeletal intermediate filaments under conditions characterized by increased cell proliferation and death.

Two layers of extra-embryonic endoderm, viz. the parietal endoderm (PE) and the visceral endoderm (VE), arise in the mouse embryo shortly after implantation. Both cell populations apparently originate from the primitive endoderm of the blastocyst. While the endoderm differentiation has been studied both in the embryo and in the embryonal carcinoma model system, the investigation has been hampered by the paucity of unequivocal markers of differentiation, especially in the case of the PE. Here we show that the PE and VE of mouse conceptuses differ in their expression of intermediate filaments: while both cell types contain cytokeratin, expression of vimentin was only revealed in the cells of the PE. The association between the differentiation of PE and the appearance of vimentin filaments is discussed.

Cytoskeleton models were produced by treating cultured human embryonal fibroblasts with Triton X-100 and actomyosin dissociating buffers. Morphological studies indicated that in addition to nuclear residues and intermediate filaments, both plasma membrane remnants and substratum-associated, pericellular material remained on the culture substratum. By immunochemical analysis the extracellular material was identified as fibronectin. The results suggest that fibronectin is associated with the nucleus-anchoring cytoskeleton of cultured human fibroblasts.

A battery of polyclonal and monoclonal antibodies directed against cytokeratins, desmin, vimentin, glial fibrillary acidic protein and neurofilament proteins of mammalian species was used to demonstrate intermediate filament (IF) expression in normal and lymphoma tissues of northern pike by indirect immunofluorescence microscopy. Frozen sections of pike intestine, skin, skeletal muscle, heart, liver, testis, head-, mid-, and posterior kidney and brain demonstrated IF in a manner which strengthens the idea that they are evolutionarily highly conserved. The results also confirm the IF-subclass specificities for different types of tissues, as noted by others in other species. Pike lymphoma cells showed morphological resemblance to head kidney cells; immunofluorescence microscopy and immunoblotting revealed vimentin expression, suggesting that the Aland pike lymphoma is a true mesenchymal neoplasm and is derived from haemic cells. The significance of these studies in relation to similar work with other species and to the possible use of IFs in the classification of normal and diseased tissues in fish is discussed.

The effects of epidermal growth factor (EGF) on the cytokeratin filaments of cultured murine epithelial cells were studied by the indirect immunofluorescence technique with affinity-purified antibodies. Mouse epithelial cells (MMC-E), grown on glass cover slips, and viewed by immunofluorescence microscopy, showed keratin-specific fluorescence as typical bright perinuclear aggregates corresponding to dense paracrystalline granules seen in electron microscopy. Within minutes after an exposure to EGF, the keratin granules in the MMC-E cells decreased. After 10 min of incubation, the cells had spread fibrillar keratin. Such an effect could not be found after a similar exposure to insulin, dexamethasone, dibutyryl cyclic AMP, or antimitotic drugs. EGF, therefore, has a relatively direct effect on the cytoskeletal organization of cultured epithelial cells. These rapid effects on the keratin filaments may explain the simultaneous EGF-induced ultrastructural surface changes of the cells. EGF may thus function as a regulatory factor in the migration of epithelial cells and in the mobility of their cell membranes. The epithelial cell line, MMC-E, should prove a useful model for studies on the action of EGF on nontransformed epithelial cells in vitro.

Three cases of pheochromocytomas (two adrenal and one retroperitoneal) were studied immunohistologically, using specific antibodies against different types of intermediate filaments. A strong positive reaction was seen in immunofluorescence microscopy with antineurofilament antibodies, while no staining of tumor cells was observed with antikeratin, antivimentin, or antidesmin antibodies. The results are in accordance with the neuroepithelial derivation of adrenal medulla and paraganglia and suggest that antineurofilament antibodies can be used as an adjunct aid in identifying pheochromocytomas.

We studied the cytoskeletal changes in natural killer (NK) cells during conjugate formation, i.e., when NK cells make contact with sensitive vs resistant target cells. F-actin and vinculin were seen to polarize at the contact sites upon conjugation with sensitive K562 cells, whereas in conjugates with resistant Raji target cells such an orientation was an infrequent finding. Myosin and two other cytoskeletal proteins, spectrin and vimentin, on the other hand, showed a random distribution in conjugating NK cells regardless of the target cell type. Hence the cytoskeletal redistribution associated with conjugation seems to be different from the receptor capping phenomenon, which is accompanied by clustering of actin, myosin, vimentin, and spectrin. On the basis of these results it seems probable that the lytic conjugate formation in NK-mediated cytotoxicity is associated with the formation of a specific type of junction that involves actin and vinculin. This cytoskeletal reorganization precedes and could be a prerequisite for the polarization of the cellular secretory apparatus and may be functionally responsible for the required cytokinetic movements.

The histogenesis of six mediastinal tumors was investigated ultrastructurally and immunohistochemically using monospecific antibodies against intermediate filament proteins. Four of the tumors, showing different appearances by light microscopy, displayed desmosomes and cytoplasmic tonofilaments, by electron microscopy, compatible with an epithelial thymoma. These cases also showed keratin positivity by immunofluorescence microscopy. One spindle cell tumor showed zonula adherens-type junctions, prominent collections of intermediate filaments, and abundant cytoplasmic neurosecretory granules consistent with a neuroendocrine tumor. In this tumor, neurofilaments could be demonstrated by immunofluorescence microscopy, a feature also consistent with a neuroendocrine tumor. One malignant tumor, lacking tonofilaments and desmosomes but showing a few primitive junctions, did not contain keratin but showed vimentin positivity. This suggests a mesenchymal origin and a diagnosis of primitive sarcoma. These cases illustrate the diagnostic usefulness of electron microscopy and immunohistochemical evaluation of intermediate filaments.

Previous studies have suggested that cytoskeletal elements in an altered form in vivo (alcoholic hyalin , AH) are targets for an autoimmune reaction in alcoholic liver disease (ALD). In this study we assayed autoantibodies to cytoskeletal cytoskeletal cytokeratin filaments, intermediate filaments of epithelial cells, by indirect immunofluorescence technique (IIF) using cultured human amnion epithelial cells and PtK 2 cells as targets. Sera from 20 patients with ALD, 38 patients with urogenital cancer, 22 patients with renal diseases, and from 18 healthy controls were studied. Patients with ALD had a significantly increased prevalence of cytokeratin filament autoantibodies using both PtK 2 cells (69%) and human amnion epithelial cells (69%) as substrate, as compared to other groups. The antibodies were mainly of IgA and IgM class and could be demonstrated in all forms of ALD. Interestingly, cytoskeleton antibodies have previously been observed in diseases of viral and "autoimmune" origin.

A 140 000 D glycoprotein (140 kD gp), labelled radioactively with surface-specific techniques, remained as the major cell surface glycoprotein in the detergent-resistant cytoskeletal preparations of cultured human fibroblasts. The 140 kD gp was present also in trypsinized cells and was not affected by treatment of the cells either with collagenase, chymotrypsin or thrombin. In density gradient fractionation of whole cells the 140 kD gp was recovered in the plasma membrane fraction together with small amounts of cytoskeletal components. In fractionation of cytoskeletal preparations, on the other hand, the 140 kD gp could not be dissociated from cytoskeletal proteins and together with vimentin it formed the major component of the oligomeric polypeptide complex generated by treating the surface-labelled cytoskeletal preparations with bifunctional cross-linking reagent, dithiobis succinimidyl propionate (DTPS). Moreover, the 140 kD gp seemed to copurify with vimentin upon reconstitution of intermediate filaments from urea-solubilized cytoskeletal preparations. On the other hand, low ionic-induced degradation of vimentin led to a decrease in the amount of the detergent-resistant 140 kD gp on the cell surface. In electron microscopy, a close apposition between bilayer-like plasma membrane remnants of the adherent cytoskeletons and cytoskeletal elements could be seen. The results indicate that the 140 kD gp is a plasma membrane glycoprotein which closely interacts with the detergent-resistant cytoskeleton of cultured human fibroblast. Possible mechanisms of the association are discussed.

Isolated neuronal perikarya from the brains of patients with Alzheimer's disease were examined in indirect immunofluorescence microscopy with different types of specific antisera against the subunit proteins of cytoskeletal intermediate filaments. From 30 to 50% of the neurofibrillary tangles were stained with antisera against each of the neurofilament triplet proteins, but not with antisera against glial fibrillary acidic protein, vimentin, desmin or cytokeratin. Polyacrylamide gel electrophoresis of Alzheimer perikaryal fractions showed polypeptide patterns markedly similar to control neuronal fractions. Our results thus suggest either that the Alzheimer neurofibrillary tangles and the antigenic neurofilament triplet protein fractions may contain related antigenic determinants or that unaffected perikaryal neurofilament material is associated with the tangles.

The complete nucleotide sequence coding for the chicken brain alpha-spectrin was determined. It comprises the entire coding frame, 5'- and 3'-untranslated sequences terminating in a poly(A)-tail. The deduced amino acid sequence shows that the alpha-chain contains 22 segments, 20 of which correspond to the typical 106 residue repeat of the human erythrocyte spectrin. Some segments non-homologous to the repeat structure reside in the middle and COOH-terminal regions. Sequence comparisons with other proteins show that these segments evidently harbour some structural and functional features such as: homology to alpha-actinin and dystrophin, two typical EF-hand structures (calcium-binding) and a putative calmodulin-binding site in the COOH-terminus and a sequence homologous to various src-tyrosine kinases and to phospholipase C in the middle of the molecule. Comparison of our sequence with other partial alpha-spectrin sequences shows that alpha-spectrin is well conserved in different species and that the human erythrocyte alpha-spectrin is divergent.

Antibodies against different cytoskeletal proteins were used to study the cytoskeletal organization of human spermatozoa. A positive staining with actin antibodies was seen in both the acrosomal cap region and the principal piece region of the tail. However, no staining was obtained with nitrobenzoxadiazol-phallacidin, suggesting that most of the actin was in the nonpolymerized form. Most of the myosin immunoreactivity was confirmed to a narrow band in the neck region of spermatozoa. Tubulin was located to the entire tail, whereas vimentin was only seen in a discrete band-like structure encircling the sperm head, apparently coinciding with the equatorial segment region. Surface staining of the spermatozoa with fluorochrome-coupled Helix pomatia agglutinin revealed a similar band-like structure that co-distributed with the vimentin-specific staining. Instead, other lectin conjugates used labeled either the acrosomal cap region (peanut and soybean agglutinins), both the acrosomal cap and the postacrosomal region of the head (concanavalin A), or the whole sperm cell surface membrane (wheat germ and lens culinaris agglutinins and ricinus communis agglutinin l). In lectin blotting experiments, the Helix pomatia agglutinin-binding was assigned to a 80,000-mol-wt polypeptide which, together with vimentin, also resisted treatment with Triton X-100. Only the acrosomal cap and the principal piece of the tail were decorated with rabbit and hydridoma antibodies against an immunoanalogue of erythrocyte alpha-spectrin (p230). p230 appeared to be the major calmodulin-binding polypeptide in spermatozoa, as shown by a direct overlay assay of electrophoretic blots of spermatozoa with 125I-calmodulin. The results indicate that spermatozoa have a highly specialized cytoskeletal organization and that the distribution of actin, spectrin, and vimentin can be correlated with distinct surface specializations of the sperm cells. This suggest that cytoskeleton may regulate the maintenance of these surface assemblies and, hence, affect the spermatozoan function.

Immunofluorescence and immunoblotting techniques were used to study the presence and distribution of vimentin and keratin type intermediate filaments, actin, and vinculin (130 kD protein) during retinoic acid (RA)-induced differentiation of F9 embryonal carcinoma (EC) cells. The undifferentiated F9 cells regularly expressed vimentin, usually concentrated close to the nucleus, but not keratin. Actin appeared as short intracellular filaments and as spikes at the edges of the colonies, together with some diffuse cytoplasmic staining. F9 cells also showed a weak, diffuse cytoplasmic vinculin-specific fluorescence in addition to occasional small focal vinculin patches at the edges of the cell colonies. RA treatment led into a series of changes in the cytoskeletal organization of F9 cells. These changes were initiated by the appearance of distinct vinculin plaques and followed by formation of actin stress fibers and by profound changes in the organization of vimentin in the flattening cells. RA treatment finally led to the appearance and co-expression of keratin fibrils in many of the vimentin-containing F9 cells. This sequence of changes suggests that the vinculin-containing adhesion plaques may be important in the mechanism of RA-induced differentiation of EC cells.

A monoclonal antibody specific for vimentin is described which, by immunofluorescence and immunoelectron microscopy, decorates fibrillar and/or granular structures in mitotic and early postmitotic cells but does not react with vimentin filaments of interphase stages of various cultured cells (rat vascular smooth muscle-derived cell line RVF-SM; SV40-transformed human fibroblasts; bovine kidney epithelial cells of line MDBK). These observations indicate that the organization of vimentin filaments varies during the cell cycle, undergoing a perimitotic change of filament organization. These changes of vimentin filaments are described in relation to those reported for cytokeratin filaments of various epithelial and carcinoma cells. The possible functional implications of filament protein rearrangements both during the cell cycle and in cell differentiation processes are discussed.

Expression of intermediate filaments was studied immunohistologically in oat-cell (6 cases), epidermoid (9 cases), adeno- (7 cases), large-cell anaplastic (3 cases), and bronchioalveolar carcinoma (3 cases), of lung. Affinity-purified antibodies against epithelial (anti-keratin), neural (anti-neurofilament), muscle (anti-desmin) and mesenchymal (anti-vimentin) intermediate filaments were used. In indirect immunofluorescence microscopy of oat-cell carcinomas a positive cytoplasmic fluorescence was seen only with antibodies against neural intermediate filaments, neurofilaments, while no staining of the tumor cells was observed with antibodies against other types of intermediate filaments. On the other hand, all the epidermoid, adeno, anaplastic, and bronchioalveolar carcinomas showed constantly a strong reaction with anti-keratin antibodies in a varying number of cells but no decoration with anti-neurofilament antibodies. The results show that expression of neural intermediate filaments is a major distinguishing feature between oat-cell carcinomas and other lung cancers and suggest that anti-neurofilament antibodies can be used as a diagnostic aid in the surgical pathologic study of pulmonary neoplasms.

Monospecific antibodies and indirect immunofluorescent microscopic examination, combined with immunochemical analysis, were used to examine intermediate filaments in four cases of bronchial carcinoid tumors. The results show that carcinoid cells express intermediate filaments of neural type (neurofilaments) but are negative for intermediate filaments of mesenchymal type (vimentin), epithelial type (keratin), muscle type (desmin), and glial type (glial fibrillary acidic protein). Since the expression of intermediate filaments shows a high degree of tissue specificity, the results suggest either derivation of bronchial carcinoid cells from maternal cells displaying neural characteristics or from cells with the capacity to acquire neural properties on neoplastic growth. It is also suggested that antineurofilament antibodies can be used as a useful aid in differential diagnosis of bronchial carcinoids from other pulmonary tumors.

We studied the effect of microtubule-disrupting drugs and cold treatment on the organization of cytoplasmic microtubules and intermediate filaments by the indirect immunofluorescence (IFL) technique. Treatment of the cultured fibroblasts with demecolcine brought about a rapid and complete disruption of microtubules and a concomitant redistribution of intermediate filaments into coiling perinuclear bundles. Cold-treatment of the cells resulted also in a complete disappearance of microtubules but did not cause any change in the distribution of intermediate filaments. During the reorganization process of the cytoplasmic microtubular complex, apparently normal arrays of intermediate filaments were discernible in the cells. The results show that microtubules can undergo a disruption-reorganization process without changes in the organization of intermediate filaments and suggest that the maintenance of normal intermediate filament organization is independent of microtubules.

In this study we characterized a skin tumor that grew in the temporal region of a 69-year-old woman. On the basis of tumor morphology, a metastasis from a small cell carcinoma of the lung was initially suggested, but X-ray and bronchoscopic studies were negative. The tumor recurred twice within a year, yet no tumors were found elsewhere in the body. Ultrastructurally, cytoplasmic organelles compatible with neuroendocrine storage granules and perinuclear aggregates of intermediate-sized (8-10 nm) filaments were found in many tumor cells. Indirect immunofluorescence microscopy revealed neurofilament-type intermediate filaments in the tumor cells but no keratin- or vimentin-type filaments. Our results further demonstrate neural properties of this tumor type, which is generally considered to have its origin from Merkel cells, the cutaneous neuroendocrine cells.

Cytoskeleton of chicken erythrocytes was studies studied after extraction of the cells with Triton X-100. In phase contrast microscopy the extracted cells were seen as ghost-like structures with preserved morphology, distinct nucleus and surrounding plasma membrane remnant. In electron microscopy, dense matrix-like nucleus, fibrillar plasma membrane residue and filaments, ca. 10 nm in diameter, traversing the cytoplasmic domain, were seen. Distinct bands of molecular weight 68000, 53000, 32000 and bands of both higher and lower molecular weight were obtained by polyacrylamide gel electrophoresis of the extracted cells. These results indicate that intermediate filaments, forming the nucleus-anchoring cytoskeleton, are present in nucleated chicken erythrocytes as part of cellular cytoskeleton.

Four cases of Ewing's sarcoma, three in bone and one from an extraskeletal site, were studied immunohistologically using monospecific antibodies against intermediate filament proteins of keratin, vimentin, desmin and neurofilament types. All cases were also evaluated for the presence of Factor VIII-related antigen (FVIIIR:Ag) and for the binding of Ulex europaeus I lectin (UEA I), both of which are endothelial markers. In all cases the tumor cells contained vimentin but not keratin, desmin or neurofilaments. The tumor cells could not be decorated with either anti-FVIIIR:Ag or UEA I, whereas the vascular endothelium was positive for both markers. The vimentin-positivity indicates a mesenchymal derivation of Ewing's sarcoma, while the lack of endothelial markers argues against the proposed endothelial origin of this tumor.

The expression of intermediate filaments of the keratin- and the vimentin-type was studied in heterokaryons of human fibroblasts and amnion epithelial cells by immunofluorescence microscopy. Fibroblasts and their homokaryons showed a fibrillar, vimentin-specific fluorescence throughout the cytoplasm but were negative when stained for keratin. Amnion epithelial cells and their homokaryons, on the other hand, showed a keratin-specific fibrillar staining, and only some of them contained also detectable vimentin. When suspended epithelial cells were fused with adherent fibroblasts, keratin fibrils spread within 3 h into the fibroblasts, intermixing with the vimentin fibrils. 1-3 d after fusion, both vimentin and keratin filaments were expressed as typical fibrillar cytoplasmic arrays, and the distribution of keratin in heterokaryons resembled closely that of vimentin. A typical cell-to-cell arrangement of keratin fibrils, seen in cultures of amnion epithelial cells, could also be found between heterokaryons. Treatment of the cultures with vinblastine sulphate induced coiling of the vimentin filaments in both homo- and heterokaryons, whereas the keratin organization was only slightly affected. Our results show that both vimentin and keratin filaments are incorporated into the cytoskeleton of heterokaryons formed between fibroblasts and epithelial cells, and that they behave in the same way as in their parental cells. Both epithelial and fibroblastic characteristics thus appear to the coexpressed in such heterokaryons.

Cultured human fibroblasts showed a typical fibrillar organization of microtubules in immunofluorescence, including the vimentin type of intermediate filament as well as actin-containing microfilaments. During infection with herpes simplex virus type 1 (HSV-1), the vimentin organization was maintained whereas actin, myosin and tubulin showed a progressive association with the viral glycoproteins within juxtanuclear structures. These structures could also be revealed with fluorochrome-coupled wheat germ agglutinin. Disruption of the microtubules by demecolcine treatment or their stabilization by taxol treatment did not prevent the aggregation of viral proteins in the cytoplasm. Taxol stabilization of the microtubules allowed the juxtanuclear accumulation of the glycoproteins in HSV-infected cells whereas treatment with demecolcine led to an accumulation of the glycoproteins either in small vesicles in the cytoplasm or in the focal adhesion areas of the cells. Production of infectious intracellular virus particles was reduced in cells treated with demecolcine or with taxol before and during infection. The results of this study indicate that the normal intracellular transport and distribution of the HSV glycoproteins and the formation of infectious virus are dependent on the presence of intact microtubules.

The presence of different types of intermediate filaments (IMF) was studied in normal ovarial tissue and in ovarian tumors. The coelomic epithelium contained both prekeratin and vimentin, but the graafian follicle cells and stromal cells contained only vimentin. In normal ovaries, desmin positivity was confined to the arterial walls. Tumors derived from surface epithelium, including Brenner tumors, expressed prekeratin in the epithelial cells, whereas sex cord-stromal tumors showed only vimentin-type IMF. Germinal tumors also showed only vimentin-type IMF, whereas differentiated teratomas expressed the IMF of the corresponding differentiation line, as recognized by light microscopy. Thus, the skin, skin appendages, and other epithelial components of dermoid cysts showed prekeratin; mesenchymal components showed vimentin (but smooth muscle cells, desmin), and glial-like areas showed glial fibrillary acidic protein. The results suggest that expression of IMF in ovarian tumors corresponds to the filament expression of the parental tissues and that the typing of IMF can be used in the differential diagnosis between ovarian tumors of different histogenesis.

Most mammalian nucleated cells contain a cytoplasmic fibril system called intermediate filaments. Unlike other cytoskeletal proteins, the subunit proteins of intermediate filaments show a remarkable cell-type-specificity in their expression: mesenchymal, muscle, epithelial, glial and neuronal cells containing each their cell type specific filaments. In this review we discuss the possibilities to use antibodies to these filaments in the diagnosis and histogenetic analysis of human tumors. This approach is based on findings which indicate that these filaments retain their cell-type specific expression also in tumors.

We studied the cytoskeletal composition of human and rat testicular myoid cells by using immunofluorescence microscopy with polyclonal and monoclonal antibodies. In adult human and rat testis, the peritubular myoid cell layer was brightly positive for desmin, the muscle type of intermediate filament protein, and a faint reaction was also seen with antibodies to vimentin, the intermediate filament protein of fibroblasts and diverse other mesenchymal cells. The desmin-positive myoid cell layer could already be identified in newborn rat testis but was more compact in appearance 23 days after birth. Both squash preparations and cultured cells from adult rat seminiferous tubules revealed distinct cell populations positive for desmin. The adult myoid cells of both species also showed a strong reaction with antibodies to myosin and p230, a nonerythroid avian alpha-spectrin analogue. The immunostaining results could be confirmed by the western blotting technique: Experiments with isolated seminiferous tubules showed a specific reaction with a 55,000-dalton and a 58,000-dalton polypeptide when desmin and vimentin antibodies were used, respectively. The present results show that the peritubular myoid cells are genuine smooth muscle cells with desmin-type intermediate filament cytoskeleton and suggest that these cells can be identified by this feature before their ultrastructural maturation.

Intermediate filaments (IF) are ubiquitous cytoplasmic structures which, by virtue of their cell- and tissue-type-specific characteristics, are widely used as markers of tissue derivation and as differential diagnostic aids in surgical pathology. In contradistinction to other IFs, vimentin filaments, characteristic of mesenchymal cells, may be co-expressed with other cell-type specific IFs--cytokeratin filaments, desmin filaments, glial filaments and neurofilaments--in some tumor cells, embryonic cells, and cells in vitro. In this study we describe a novel type of IF co-expression which does not involve vimentin-filaments, viz. the presence of both cytokeratin filaments and neurofilaments in human bronchial carcinoid tumor cells.

The cytoskeleton of human ventricular myocardium in normal and pathological hearts has been analysed with immunocytochemical techniques. Specific antibodies against the intermediate filament proteins desmin (Mr 55000) and vimentin (Mr 58000) and antibodies against two cytoskeleton-associated proteins, a spectrin-like protein (Mr 230 000) and vinculin (Mr 130000), have been used. We show that desmin is localized in the myocytes as an intermyofibrillar lattice at the Z disk level of the myofibrils, and at the intercalated disks. The spectrin-like protein is localized as a transverse striated pattern interlinked with fine longitudinal strands in the subplasmalemmal region of the myocytes. Vinculin is abundant in the intercalated disks and in myotendinous junctions but occurs also at the peripheral sarcolemma in the form of a regular repeat of dots and of fine bar-like extensions into the cytoplasm from the dots. These patterns were observed both in normal and in abnormal hearts, but a number of altered patterns in pathological myocytes were also seen. It is concluded that the intermediate filament system has important implications in the structural function of normal and abnormal hearts but that further studies are needed to elucidate how the different components are related to each other and how they are influenced by different disease processes. [ABSTRACT FROM PUBLISHER]