Live stripping of clathrin-coated vesicles.

Vesicle budding requires recruitment of a coat, which must then be removed to allow fusion with the target compartment. In vitro assays have implicated Hsc70 and auxilin family members as key players in clathrin-coated vesicle uncoating. New in vivo studies now show that this is indeed the case and reveal additional functions of the Hsc70/auxilin complex.

Polarized expression of integrin receptors (alpha 6 beta 4, alpha 2 beta 1, alpha 3 beta 1, and alpha v beta 5) and their relationship with the cytoskeleton and basement membrane matrix in cultured human keratinocytes.

In human keratinocytes cultured in conditions which allow differentiation and stratification and are suitable to reconstitute a fully functional epidermis, alpha 6 beta 4 and two members of the beta 1 integrin family (alpha 2 beta 1 and alpha 3 beta 1) were respectively polarized to the basal and lateral domains of the plasmamembrane both in growing colonies and in the reconstituted epidermis. Conversely, the alpha v integrin subunit, presumably in association with beta 5, was expressed at the basal surface in growing and migrating but not in stationary keratinocytes. The integrin alpha 6 beta 4: (a) was organized in typical patches which often showed a "leopard skin" pattern where spots corresponded to microfilament-free areas; (b) was not associated with focal contacts containing vinculin and talin but rather corresponded to relatively removed contact areas of the basal membrane as shown by interference reflection microscopy; and (c) was coherent to patches of laminin secreted and deposited underneath the ventral membrane of individual cells. The two beta 1 integrins (alpha 2 beta 1 and alpha 3 beta 1), both endowed with laminin receptor properties, were not associated with focal adhesions under experimental conditions allowing full epidermal maturation but matched the lateral position of vinculin (but not talin), cingulin, and desmoplakin, all makers of intercellular junctions. Often thin strips of laminin were observed in between the lateral aspects of individual basal keratinocytes. The integrin complex alpha v beta 5 had a topography similar to that of talin- and vinculin-containing focal adhesions mostly in the peripheral cells of expanding keratinocyte colonies and in coincidence with fibronectin strands. The discrete topography of beta 1 and beta 4 integrins has a functional role in the maintenance of the state of aggregation of cultured keratinocytes since lateral aggregation was impaired by antibodies to beta 1 whereas antibodies to beta 4 prevented cell-matrix adhesion (De Luca, M., R. N. Tamura, S. Kajiji, S. Bondanza, P. Rossino, R. Cancedda, P. C. Marchisio, and V. Quaranta. Proc. Natl. Acad. Sci. USA. 87:6888-6892). Moreover, the surface polarization of integrins followed attachment and depended both on the presence of Ca2+ in the medium and on the integrity of the cytoskeleton. We conclude that our in vitro functional tests and structural data suggest a correlation between the pattern of integrin expression on defined plasmamembrane domains and the mechanism of epidermal assembly.

Amphiphysin II (SH3P9; BIN1), a member of the amphiphysin/Rvs family, is concentrated in the cortical cytomatrix of axon initial segments and nodes of ranvier in brain and around T tubules in skeletal muscle.

Amphiphysin (amphiphysin I), a dominant autoantigen in paraneoplastic Stiff-man syndrome, is a neuronal protein highly concentrated in nerve terminals, where it has a putative role in endocytosis. The yeast homologue of amphiphysin, Rvs167, has pleiotropic functions, including a role in endocytosis and in actin dynamics, suggesting that amphiphysin may also be implicated in the function of the presynaptic actin cytoskeleton. We report here the characterization of a second mammalian amphiphysin gene, amphiphysin II (SH3P9; BIN1), which encodes products primarily expressed in skeletal muscle and brain, as differentially spliced isoforms. In skeletal muscle, amphiphysin II is concentrated around T tubules, while in brain it is concentrated in the cytomatrix beneath the plasmamembrane of axon initial segments and nodes of Ranvier. In both these locations, amphiphysin II is colocalized with splice variants of ankyrin3 (ankyrinG), a component of the actin cytomatrix. In the same regions, the presence of clathrin has been reported. These findings support the hypothesis that, even in mammalian cells, amphiphysin/Rvs family members have a role both in endocytosis and in actin function and suggest that distinct amphiphysin isoforms contribute to define distinct domains of the cortical cytoplasm. Since amphiphysin II (BIN1) was reported to interact with Myc, it may also be implicated in a signaling pathway linking the cortical cytoplasm to nuclear function.

alpha-2 Macroglobulin receptor/Ldl receptor-related protein(Lrp)-dependent internalization of the urokinase receptor.

The GPI-anchored urokinase plasminogen activator receptor (uPAR) does not internalize free urokinase (uPA). On the contrary, uPAR-bound complexes of uPA with its serpin inhibitors PAI-1 (plasminogen activator inhibitor type-1) or PN-1 (protease nexin-1) are readily internalized in several cell types. Here we address the question whether uPAR is internalized as well upon binding of uPA-serpin complexes. Both LB6 clone 19 cells, a mouse cell line transfected with the human uPAR cDNA, and the human U937 monocytic cell line, express in addition to uPAR also the endocytic alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein (LRP/alpha 2-MR) which is required to internalize uPAR-bound uPA-PAI-1 and uPA-PN-1 complexes. Downregulation of cell surface uPAR molecules in U937 cells was detected by cytofluorimetric analysis after uPA-PAI-1 and uPA-PN-1 incubation for 30 min at 37 degrees C; this effect was blocked by preincubation with the ligand of LRP/alpha 2-MR, RAP (LRP/alpha 2-MR-associated protein), known to block the binding of the uPA complexes to LRP/alpha 2-. MR. Downregulation correlated in time with the intracellular appearance of uPAR as assessed by confocal microscopy and immuno-electron microscopy. After 30 min incubation with uPA-PAI-1 or uPA-PN-1 (but not with free uPA), confocal microscopy showed that uPAR staining in permeabilized LB6 clone 19 cells moved from a mostly surface associated to a largely perinuclear position. This effect was inhibited by the LRP/alpha 2-MR RAP. Perinuclear uPAR did not represent newly synthesized nor a preexisting intracellular pool of uPAR, since this fluorescence pattern was not modified by treatment with the protein synthesis inhibitor cycloheximide, and since in LB6 clone 19 cells all of uPAR was expressed on the cell surface. Immuno-electron microscopy confirmed the plasma membrane to intracellular translocation of uPAR, and its dependence on LRP/alpha 2-MR in LB6 clone 19 cells only after binding to the uPA-PAI-1 complex. After 30 min incubation at 37 degrees C with uPA-PAI-1, 93% of the specific immunogold particles were present in cytoplasmic vacuoles vs 17.6% in the case of DFP-uPA. We conclude therefore that in the process of uPA-serpin internalization, uPAR itself is internalized, and that internalization requires the LRP/alpha 2-MR.

Expression, topography, and function of integrin receptors are severely altered in keratinocytes from involved and uninvolved psoriatic skin.

Psoriasis is a hyperproliferative cutaneous disease of unknown etiology and etiopathogenesis. Alteration of keratinocyte adhesiveness to basal lamina has been proposed as the initial disturbance leading to poorly controlled proliferation. Keratinocyte adhesion to basal lamina and lateral interactions among basal epidermal cells are mediated, besides other molecules, by integrin receptors that are segregated to discrete membrane domains. In this paper, the expression and function of integrins in psoriatic keratinocytes were examined, both in vivo and in vitro. We found that: (a) in psoriatic keratinocytes the integrin heterodimers alpha 2 beta 1, alpha 3 beta 1, and alpha 6 beta 4 have lost their polarized distribution on the plasma membrane; (b) the role of these integrins in mediating keratinocyte adhesion in vitro is altered; (c) psoriatic keratinocytes form focal contacts containing both beta 1 and beta 4 integrins. In normal adult keratinocytes the alpha 5 beta 1 fibronectin receptor is poorly expressed and diffusely distributed on the basal keratinocyte plasma membrane and is not organized in defined adhesive structures. In contrast, psoriatic keratinocytes show a clear fibronectin receptor staining in vivo, and organize alpha 5 beta 1 in typical focal contacts in vitro without any obvious increase of its expression and synthesis. These multiple alterations of integrins are also present in uninvolved keratinocytes from psoriatic patients, suggesting a key role for altered integrin-mediated adhesion in the pathogenesis of this disease.

In vitro and in vivo activation of endothelial cells by colony-stimulating factors.

This study was designed to identify the set of functions activated in cultured endothelial cells by the hematopoietic growth factors, granulocyte colony-stimulating factor (G-CSF) and granulocyte macrophage-colony-stimulating factor (GM-CSF), and to compare them with those elicited by prototypic cytokines active on these cells. Moreover, indications as to the in vivo relevance of in vitro effects were obtained. G-CSF and GM-CSF induced endothelial cells to proliferate and migrate. In contrast, unlike appropriate reference cytokines (IL-1 and tumor necrosis factor, IFN-gamma), G-CSF and GM-CSF did not modulate endothelial cell functions related to hemostasis-thrombosis (production of procoagulant activity and of platelet activating factor), inflammation (expression of leukocyte adhesion molecule-1 and production of platelet activating factor), and accessory function (expression of class II antigens of MHC). Other colony-stimulating factors (IL-3 and macrophage-colony-stimulating factor) were inactive on all functions tested. In comparison to basic fibroblast growth factor (bFGF), G-CSF and GM-CSF induced lower maximal proliferation of endothelial cells, whereas migration was of the same order of magnitude. G-CSF and GM-CSF stimulated repair of mechanically wounded endothelial monolayers. Exposure to both cytokines induced shape changes and cytoskeletal reorganization consistent with a migratory phenotype. To explore the in vivo relevance of the in vitro effects of these cytokines on endothelium, we studied the angiogenic activity of human G-CSF in the rabbit cornea. G-CSF, but not the heat-inactivated molecule, had definite angiogenic activity, without any sign of inflammatory reactions. G-CSF was less active than bFGF. However, the combination of a nonangiogenic dose of bFGF with G-CSF resulted in an angiogenic response higher than that elicited by either individual cytokines. Thus, G-CSF and GM-CSF induce endothelial cells to express an activation/differentiation program (including proliferation and migration) related to angiogenesis.

Synaptic vesicle endocytosis.

Exocytosis of synaptic vesicles is followed rapidly by reinternalization and recycling of their membranes. Recent studies have confirmed the key role of clathrin-mediated endocytosis in synaptic vesicle reformation and have identified new proteins that participate in this process. In addition, growing evidence suggests that lipids, primarily phosphoinositides, play an important role in synaptic vesicle recycling.

Changes in integrin and E-cadherin expression in neoplastic versus normal thyroid tissue.

UNLABELLED: BACK: The functional organization of polarized epithelia depends mostly on adhesion molecules belonging to the integrin and cadherin families. These molecules either recognize basement membrane components, such as laminins, or form intercellular junctions via homotypic interactions. Such tissue organization is often disrupted upon neoplastic transformation, and the resulting loss of functional polarization and cell cohesion might be a prerequisite for the invasive and metastatic behavior of carcinomas. PURPOSE: We studied modifications on thyroid adhesive mechanisms at various stages of neoplastic progression in terms of adhesion molecule expression, topography, and functional regulation by tyrosine kinases. Starting from this working hypothesis, we sought to identify one or more biological markers that would be suggestive of malignant transformation and poorer prognosis and that could be developed as a reliable indicator(s) in early diagnostic steps. METHODS: The study was carried out on both surgical samples and the corresponding fine-needle aspiration biopsy smears (numbers of specimens collected: 19 adenomas, seven follicular carcinomas, 13 papilary carcinomas, and 39 normal tissues). Immunohistochemistry of tissue sections and smears and immuno-precipitation and western blot analysis of protein extracts were done with a battery of monoclonal and polyclonal antibodies. Northern blotting was performed on RNA extracts from frozen tissue samples and use of an integrin subunit beta4 complementary DNA probe. RESULTS: Our findings can be summarized as follows: 1) In normal thyroid cells, the cooperative role of integrin alpha6beta4 and laminin 5/kalinin in hemidesmosome-mediated adhesion adhesion is missing, and recognition of the basal lamina occurs via integrin alpha3beta1 and laminin 1 and/or 2 (this pattern being maintained in adenomas but altered in carcinomas regardless of their histotype or differentiation grade); 2) only in carcinomas with clinical and/or histologic aggressiveness do neoexpression of integrin subunit beta4 and loss of laminin 2/merosin occur, indicating de novo assembly of integrin alpha6beta4; 3) pericellular redistribution and cytoskeletal disconnection of the E-cadherin-catenin complex occur; and 4) basal E-cadherin tyrosine phosphorylation decreases in carcinomas as compared with that in normal and adenomatous tissue. CONCLUSION: The malignant progression of thyroid tumors involves marked rearrangement of cell-basement membrane and cell-cell adhesion molecules and changes in their cytoskeleton linkage. These rearrangements are also easily and reproducibly detected on fine-needle aspiration biopsy smears. IMPLICATIONS: Immunodetection of adhesion molecules in sections and/or fine-needle smears may complement the toolbox of thyroid surgical pathologists; it may expand the possibilities of achieving a correct early diagnosis of thyroid tumors and of gaining some prognostic information on thyroid tumors.

The Met/HGF receptor is over-expressed in human osteosarcomas and is activated by either a paracrine or an autocrine circuit.

The c-MET oncogene encodes the receptor for the Hepatocyte Growth Factor/Scatter Factor (HGF), a cytokine that stimulates the invasive growth of normal and neoplastic cells. The Met/HGF receptor is expressed by epithelial cells and its ligand by cells of mesenchymal origin. Receptor-ligand interaction occurs via a paracrine circuit. We studied the expression of the Met/HGF receptor and of its ligand in mesenchymal human tumours by examining 39 clinical samples of bone tumours. The Met/HGF receptor was not detectable in the majority of bone tumours, as expected from their mesenchymal origin. Notably, the receptor was overexpressed in 60% of the osteosarcomas examined. In 12 osteosarcoma cell lines the Met/HGF receptor was overexpressed, phosphorylated by HGF stimulation and fully functional. HGF was detected in two out of seven clinical specimens of osteosarcoma. The ligand and the receptor are co-expressed in two clonal osteosarcoma cell lines. In these lines the Met/HGF receptor was constitutively phosphorylated; phosphorylation was suppressed by suramin treatment, a known blocker of autocrine loops. These data suggest that activation of the Met/HGF receptor by a paracrine or an autocrine mechanism might play a role in the particularly aggressive behaviour of osteosarcomas.

Synaptojanin 1 contributes to maintaining the stability of GABAergic transmission in primary cultures of cortical neurons.

Inhibitory synapses in the CNS can exhibit a considerable stability of neurotransmission over prolonged periods of high-frequency stimulation. Previously, we showed that synaptojanin 1 (SJ1), a presynaptic polyphosphoinositide phosphatase, is required for normal synaptic vesicle recycling (Cremona et al., 1999). We asked whether the stability of inhibitory synaptic responses was dependent on SJ1. Whole-cell patch-clamp recordings of unitary IPSCs were obtained in primary cortical cultures between cell pairs containing a presynaptic, fast-spiking inhibitory neuron (33.5-35 degrees C). Prolonged presynaptic stimulation (1000 stimuli, 2-20 Hz) evoked postsynaptic responses that decreased in size with a bi-exponential time course. A fast component developed within a few stimuli and was quantified with paired-pulse protocols. Paired-pulse depression (PPD) appeared to be independent of previous GABA release at intervals of >/=100 msec. The characteristics of PPD, and synaptic depression induced within the first approximately 80 stimuli in the trains, were unaltered in SJ1-deficient inhibitory synapses. A slow component of depression developed within hundreds of stimuli, and steady-state depression showed a sigmoidal dependence on stimulation frequency, with half-maximal depression at 6.0 +/- 0.5 Hz. Slow depression was increased when release probability was augmented, and there was a small negative correlation between consecutive synaptic amplitudes during steady-state depression, consistent with a presynaptic depletion process. Slow depression was increased in SJ1-deficient synapses, with half-maximal depression at 3.3 +/- 0.9 Hz, and the recovery was retarded approximately 3.6-fold. Our studies establish a link between a distinct kinetic component of physiologically monitored synaptic depression and a molecular modification known to affect synaptic vesicle reformation.

Integrin distribution and cytoskeleton organization in normal and malignant monocytes.

In this work we have mapped by double-label immunofluorescence the cellular distribution of integrins and their relationship with cytoskeletal proteins in normal and malignant monocytes. In normal monocytes, CD18 and CD11c are concentrated at specific adhesion sites, named podosomes, together with actin, vinculin, and talin, while CD11a, CD11b, CD29/beta 1, CDw49d/alpha 4 and CD54/ICAM-1 retain a diffuse distribution on the cell surface without a selective pattern of localization. U-937 and fresh leukemic monoblasts under standard culture conditions do not adhere and do not form podosomes, but, when treated with TPA, they promptly adhere to substrate, form podosomes and focal adhesions in different cells and display the same integrin/cytoskeleton relationship as normal mature monocytes. Further, in these cells CD18, CD11a, CD11c, ICAM-1, and talin, but not vinculin, co-localize in homotypic cell junctions, thus showing a close relationship between integrins and talin. These observations provide morphological evidence that, in cells of the monocytic lineage, podosome formation is acquired upon differentiation and different integrins are selectively localized at different adhesion sites.

Osteoclast precursors circulate in the peripheral blood of patients with aggressive multiple myeloma.

Osteolysis resulting in extensive bone damage is a major clinical manifestation of patients with multiple myeloma (MM). The mechanisms of bone resorption in MM are incompletely understood. The final pathway is the generation of activated osteoclasts within bone marrow (BM) microenvironment. To investigate the mechanisms of bone resorption in MM we established an experimental system that, including bone marrow (BM) stromal cells and bone slices, closely mimicks in vitro the in vivo BM microenvironment. Peripheral blood mononuclear cells (PBMC) from nine patients with MM, three monoclonal gammopathy of undetermined significance (MGUS), and nine normal controls were cultured in this system. PBMC from patients with aggressive and bone devastating MM gave rise to multi-nucleated cells with the morphology and phenotype of osteoclasts. These cells induced bone resorption in vitro which was inhibited by the addition of calcitonin. No bone resorption was observed in cultures of PBMC from patients with MM and limited bone damage, with MGUS and from normal subjects. These findings indicate that patients with aggressive MM have a population of circulating precursors that develop into functionally active osteoclast-like cells once they come in contact with the BM microenvironment. These cells may contribute to the wide-spread and generalized bone erosion observed in the patients.

Dynamics of secretion.

In this short review, kinetic aspects of exocytosis are discussed. A special emphasis is put on recent data that highlight dynamic differences between neurotransmission and other forms of secretion.

Expression and topography of integrins and basement membrane proteins in epidermal carcinomas: basal but not squamous cell carcinomas display loss of alpha 6 beta 4 and BM-600/nicein.

The expression and topography of some integrins and basement membrane proteins in cutaneous basal cell carcinomas (BCCs) and squamous cell carcinomas (SCCs) have been studied by immunohistochemistry and Western blotting. It has been shown that the typical cell-to-cell distribution of alpha 2 beta 1 and alpha 3 beta 1 found in normal epidermis is replaced by pericellular distribution in both BCC and SCC cells. BCC and SCC also showed different patterns of expression of alpha 6 beta 4, an integrin heterodimer normally lining the basal surface of basal epidermal keratinocytes: whereas SCC showed high expression and pericellular distribution of alpha 6 beta 4, BCC cells did not express this integrin at all. The absence of alpha 6 and beta 4 subunits from BCC extracts was confirmed by Western blotting. The molecular composition of the basement membrane was markedly different in the two types of epidermal tumors. Whereas laminin and collagen type IV were conserved in the basement membrane zone of both tumors, the molecular complex BM-600/nicein, which is recognized by the monoclonal antibody GB3 and is possibly identical to the previously described basement membrane glycoproteins kalinin and epiligrin, was absent from BCC cells. Then, the simultaneous loss of expression of alpha 6 beta 4 and BM-600/nicein in BCC cells but not in SCC cells indicates that alpha 6 beta 4 integrin and one of its potential ligands may be co-regulated in both BCC and SCC, thus suggesting a role for this phenomenon in the pathogenesis and clinical behavior of these epidermal tumors.

Expression and role of integrin receptors in Sézary syndrome.

The potential role of integrins in the epidermotropism of the atypical lymphocytes of Sézary syndrome was studied by monitoring the expression of alpha and beta chains and their major ligands in skin biopsies and peripheral blood cells in patients at different progression stages. Most mononuclear cell integrins were also detected on infiltrating cells including the leukocyte complex CD11/CD18, alpha 4 beta 1, and their ligands, ICAM-1 and VCAM-1. Conversely, alpha 6 and beta 4 were present only in epidermal basal cells. Mononuclear infiltrates of SS were positive for both alpha 3 and alpha 5 chains, whereas in inflammatory cutaneous diseases only alpha 5 was expressed, indicating that a major feature of Sézary cells is the unique expression of alpha 3 beta 1. Significant changes of alpha 3 beta 1 were monitored in the follow-up of Sézary patients and correlated with the results of the therapy. The heterodimer alpha 1 beta 1 was absent from mononuclear cells except in one case. Among matrix molecules, laminin and type IV collagen displayed a pattern similar to that of the controls, whereas fibronectin and tenascin deposition were apparently increased. Circulating Sézary cells, both at diagnosis and during follow-up, were alpha 3 and alpha 5 negative and failed to acquire these adhesion molecules after mitogenic stimulation. We propose that the expression of alpha 3 beta 1 is a distinguishing feature of skin-infiltrating Sézary cells and may be related to their epidermotropism. It could also be adopted as an additional parameter of the progression and therapeutic stage of Sézary syndrome.

Biosynthesis and apical localization of the urokinase receptor in polarized MDCK epithelial cells.

The biosynthesis and the surface localization of the urokinase plasminogen activator receptor (uPAR) were analysed in MDCK epithelial cells and in unpolarized fibroblasts. No differences were observed with respect to rate of synthesis, nature of precursors and time of surface appearance. uPAR was localized particularly at the focal and cell-cell contacts when expressed in fibroblasts. On the contrary, in MDCK cells uPAR was found mostly on the apical surface; in agreement with its localization, down-regulation of uPAR by the uPA-PAI-1 complex was observed only from the apical membrane.

The alpha 6 and beta 4 integrin subunits are expressed by smooth muscle cells of human small vessels: a new localization in mesenchymal cells.

The alpha 6 beta 4 integrin complex is generally thought to be expressed by epithelial cells, where it is localized in specific adhesion structures, the hemidesmosomes. Recent observations have suggested a new localization of the beta 4 integrin chain in small vessels, possibly in endothelial cells, i.e., in cells of mesenchymal origin. In the present study we show that (a) the alpha 6 and beta 4 integrin chains are not expressed by endothelial cells, since they are not localized in von Willebrand factor-producing cells; (b) instead, smooth muscle cells of small vessels are intensely positive to antibodies to both alpha 6 and beta 4 intergrin chains; and (c) in some restricted regions of these smooth muscle cells there is a clear colocalization between alpha-smooth muscle actin and alpha 6 and beta 4 integrin chains, suggesting that a new type of cytoskeletal linkage for the alpha 6 beta 4 integrin complex may occur in mesenchyme-derived cells. Our observations are supported by confocal laser microscopy (CLSM) images of specimens labeled by double immunofluorescence. This technical choice was made to take advantage of the higher resolution offered by CLSM in comparison with conventional immunofluorescence. A careful selection of barrier filters was necessary to separate accurately emission and excitation spectra of the fluorochromes used in this study, resulting in an efficient colocalization analysis.

Integrins and basement membrane proteins in skin carcinomas.

The topography of some integrins and basement membrane proteins in cutaneous basal cell carcinomas (BCC) and squamous cell carcinomas (SCC) has been studied by immunohistochemistry. The typical cell-to-cell distribution of alpha 2 beta 1 and alpha 3 beta 1 of normal epidermis becomes pericellular in both BCC and SCC cells. BCC and SCC also showed different patterns of expression of alpha 6 beta 4, the integrin normally located at the basal surface of basal epidermal keratinocytes. SCC showed high expression and pericellular distribution of alpha 6 beta 4, while BCC did not express alpha 6 beta 4 at all also in Western blotting. The basement membrane was markedly different in the two tumor types. Laminin and collagen type IV were conserved in the basement membrane zone of both tumors while nicein/kalinin was absent from BCC cells. The simultaneous loss of expression of alpha 6 beta 4 and nicein/kalinin in BCC cells but not in SCC cells indicates that alpha 6 beta 4 and one potential ligand may be co-regulated in both BCC and SCC. We suggest a role for this phenomenon in the pathogenesis of these epidermal tumors.

Distribution of integrins and extracellular matrix proteins in vulvar squamous cell carcinomas.

We report the topography of integrins and some basement membrane zone (BMZ) proteins in normal vulvar skin and in seven cases of vulvar squamous carcinoma (VSC). In vulvar epidermis integrin chains alpha 2, alpha 3 and beta 1 lined the lateral surface of basal cells, while the heterodimer alpha 6 beta 4 was detected only at their basal domain. The location of alpha 6 beta 4 exactly matched BMZ identified by kalinin, laminin and collagen type IV. In VSC this pattern was subverted since both beta 1 integrins and alpha 6 beta 4 became pericellular. In particular, alpha 6 beta 4 lost its coherence with the residual organization of BMZ. In fact, BMZ components displayed their normal pattern were this was preserved, and so were fibronectin and tenascin in the stroma underlying the tumor. Furthermore, alpha 5 beta 1, the prototype fibronectin receptor that is not normally exposed in vulvar epidermal cells, became detectable pericellularly in VSC tumor cells. Alteration of integrin polarized topography appears to be typical of VSC cells as well as of other tumor epidermal cells. This alteration may be easily shown by immunohistochemistry on routinely collected frozen biopsies. It may then represent a tool for the early diagnosis of VSC that provides a quick and easy complement of traditional histology.