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PubMed1997

Phenotypic characterization of keratinocytes migrated from polymer support - in vitro study.

The keratinocytes are able to migrate from the poly (2- hydroxyethylmethacrylate) disc if it is transferred to the new Petri-dish colonized with irradiated 3T3 mouse fibroblasts, and form a ring-shaped colony around the disc. The phenotypic characterization of human keratinocytes migrated from these discs was studied using a group of monoclonal antibodies. The keratinocytes in the external periphery of the colony of cells which migrated from the disc express the proliferating cell nuclear antigen (PCNA), alpha2, alpha3 chains and alpha5beta1 integrin receptor. A protein of the desmosome complex, desmoplakin-1, was also expressed. Involucrin and cytokeratin-10 were expressed after prolonged cultivation. These results suggest that the migrated keratinocytes are able to proliferate, recognize extracellular matrix molecules important in the process of the re-epitelization of the wound, and terminally differentiate in vitro. They are encouraging for further experiments with respect to the development of a support for keratinocyte cultivation and for grafting in clinical practice.

Journal Article↗

PubMed2001

Synthetic peptides inhibit adhesion of human tumor cells to extracellular matrix proteins.

Human tumor cell progression and metastasis are partially dependent on the ability of a tumor cell to adhere to the proteins of the extracellular matrix (ECM) and survive at the distant location. Six novel D-amino acid-containing peptides were analyzed for their ability to adhere to human prostate tumor cells, support tumor cell adhesion, and inhibit tumor cell adhesion to ECM proteins or human dermal fibroblasts. Of these, two peptides called RZ-3 (kmviywkag) and HYD-1 (kikmviswkg) bound to tumor cell surfaces and compared favorably with the previously reported AG-73 (RKRLQVQLSIRT) L-amino acid peptide, as determined by fluorescence-activated cell sorting analysis. A scrambled peptide derivative of HYD-1, called HYDS-1 (wiksmkivkg), was not active. The RZ-3, HYD-1, and AG-73 peptides supported maximal cancer cell adhesion at 5 microg, 10 microg, and 50 microg/well, respectively. The ECM proteins fibronectin, laminin 1, and collagen IV supported maximal cell adhesion at 1 microg, >10 microg, and 50 microg/well, respectively. Prostate tumor cell adhesion to immobilized RZ-3 and HYD-1 peptides was inhibited by alpha2-6- and beta1-integrin-blocking antibodies. Conversely, tumor cell adhesion to a beta1-integrin-specific antibody was blocked by both RZ-3 and HYD-1. Epithelial cell adhesion to dermal fibroblasts was inhibited by HYD-1 and unaffected by the scrambled peptide, HYDS-1. Cell adhesion to immobilized peptides was unaffected by EDTA. The soluble RZ-3 and HYD-1 peptides inhibited tumor cell adhesion to each of the immobilized four ECM proteins (1.0 microg/well) in a time- and concentration-dependent manner. The IC(50) of the RZ-3 peptide for blocking adhesion to fibronectin, laminin 1, laminin 5, and collagen IV was 2.4 microg, 1.8 microg, 4.6 microg, and 2.8 microg/well, respectively. The IC(50) of the HYD-1 peptide for blocking adhesion to fibronectin, laminin 1, laminin 5, and collagen IV was 6.9 microg, 5.7 microg, >10 microg, and 6.2 microg/well, respectively. Taken together, these results indicate that RZ-3 and HYD-1 are biologically active D-amino acid-containing peptides that can themselves support tumor cell adhesion and can inhibit tumor cell adhesion to immobilized ECM proteins or dermal fibroblasts.

Carcinoma, Squamous Cell↗

PubMed1998

Initiation of skin basement membrane formation at the epidermo-dermal interface involves assembly of laminins through binding to cell membrane receptors.

To study the mechanism of basement membrane formation, we determined by immunochemistry temporal and spatial expression of laminin-5 (Ln-5), laminin-1 (Ln-1) and their integrin receptors during early skin morphogenesis. A 3-dimensional skin culture was used that allows the study of the sequential molecular events of basement membrane formation at the epidermodermal interface. During early anchorage of keratinocytes to the extracellular matrix there is expression of Ln-5, BP-230 antigen and alpha3, beta1 integrin subunits. During epidermal stratification and prior to the formation of the lamina densa there is assembly of Ln-5, Ln-1, collagen IV and nidogen accompanied by keratinocyte basal clustering of alpha2, alpha3, alpha6, beta1, and beta4+ integrin subunits. The assembly pattern of Ln-1 and Ln-5 can be disturbed with functional antibodies against the beta1 (AIIB2) and alpha6 (GoH3) integrin subunits. Ln-1 assembly can also be disturbed with antibodies against its E8 domain and by competitive inhibition with a synthetic peptide (AG-73) derived from its G-4 domain. Quantitative RT-PCR showed that the dermis contributes about 80% of the laminin gamma)1 chain mRNA while 20% is produced by the epidermis which emphasizes its dual tissue origin and the major contribution of the mesenchyma in laminin production. The laminin gamma2 chain mRNA, present in Ln-5, was mostly of epidermal origin. This study presents evidence that during the initiation of basement membrane formation, laminins bind to keratinocyte plasma membrane receptors and thus may serve as nucleation sites for further polymerization of these compounds by a self-assembly process.

Actin Cytoskeleton↗

PubMed2006

In vitro analysis of integrin expression during chondrogenic differentiation of mesenchymal stem cells and chondrocytes upon dedifferentiation in cell culture.

Tissue engineering represents a promising method for generating chondrogenic grafts for reconstructive surgery. In cultured chondrocytes, the dedifferentiation of cells seems unavoidable for multiplication. Stem cells, however, displaying unlimited self-renewal and the capacity to differentiate towards chondrocytes, might be usable after further characterization. As the interactions between the extracellular matrix and the cellular compartment can alter the cellular behaviour, we investigated the expression of integrins using microarray analysis during chondrogenic differentiation of human mesenchymal stem cells (MSC) in comparison with de-differentiating human chondrocytes (HC) harvested during septoplasty. During chondrogenic differentiation of MSC, the fibronectin-receptor (Integrin beta1alpha5), fibronectin and the GPIIb/IIIa-receptor were downregulated. The components of the vitronectin-receptor (Integrin alphavbeta3) and CD47 were constantly expressed and ILK was downregulated. Vitronectin and osteopontin were not expressed by the cells. In HC, Integrin beta1alpha5 in conjunction with the ligand fibronectin were upregulated during dedifferentiation, Integrin alphavbeta3 as well as the GBIIb/IIIa-receptor were activated on day 21 but neither vitronectin nor osteopontin were expressed by the cells. The integrins, beta2, beta4, beta6, beta8 and alpha2, alpha4, alpha6, alpha7, alpha11, were not expressed at any time. ILK, CD47, and ICAP were activated with ongoing dedifferentiation. In conclusion, a candidate for signal-transmission is the fibronectin receptor (integrin alpha5beta1) in conjunction with its ligand fibronectin. Other receptors, e.g. for vitronectin and osteopontin (alphavbeta3), or their ligands do not seem to be involved in signal transmission for dedifferentiation. The GPIIb/IIIa-receptor might assist the process of dedifferentiation. Intracellularly, ILK, ICAP1 and CD47 might be involved in the transduction of integrin-dependent signals.

Cell Differentiation↗

PubMed1999

In vitro modulation of keratinocyte wound healing integrins by zinc, copper and manganese.

Although the trace elements zinc, copper and manganese are used in vivo for their healing properties, their mechanism of action is still only partially known. Some integrins expressed by basal layer keratinocytes play an essential part in healing, notably alpha2beta1, alpha3beta1, alpha6beta4 and alphaVbeta5, whose expression and distribution in epidermis are modified during the re-epithelialization phase. This study demonstrates how the expression of these integrins are modulated in vitro by trace elements. Integrin expression was studied in proliferating keratinocytes in monolayer cultures and in reconstituted skin that included a differentiation state. After 48 h incubation with zinc gluconate (0.9, 1.8 and 3.6 microg/mL), copper gluconate (1, 2 and 4 microg/mL), manganese gluconate (0.5, 1 and 2 microg/mL) and control medium, integrin expression was evaluated by FACScan and immunohistochemistry. Induction of alpha2, alpha3, alphaV and alpha6 was produced by zinc gluconate 1.8 microg/mL in monolayers, of alpha2, alpha6 and beta1 by copper gluconate 2 and 4 microg/mL and of all the integrins studied except alpha3 by manganese gluconate 1 microg/mL. Thus, alpha6 expression was induced by all three trace elements. The inductive effect of zinc was particularly notable on integrins affecting cellular mobility in the proliferation phase of wound healing (alpha3, alpha6, alphaV) and that of copper on integrins expressed by suprabasally differentiated keratinocytes during the final healing phase (alpha2, beta1 and alpha6), while manganese had a mixed effect.

Cells, Cultured↗

PubMed2005

Hypersialylation of beta1 integrins, observed in colon adenocarcinoma, may contribute to cancer progression by up-regulating cell motility.

Colon adenocarcinomas are known to express elevated levels of alpha2-6 sialylation and increased activity of ST6Gal-I, the Golgi glycosyltransferase that creates alpha2-6 linkages. Elevated ST6Gal-I positively correlates with metastasis and poor survival, and therefore ST6Gal-I-mediated hypersialylation likely plays a role in colorectal tumor invasion. Previously we found that oncogenic ras (present in roughly 50% of colon adenocarcinomas) up-regulates ST6Gal-I and, in turn, increases sialylation of beta1 integrin adhesion receptors in colon epithelial cells. However, we wanted to know if this pattern held true in vivo and, if so, how beta1 hypersialylation might contribute to colon tumor progression. In the present study, we find that beta1 integrins from colon adenocarcinomas consistently carry higher levels of alpha2-6 sialic acid. To explore the effects of increased alpha2-6 sialylation on beta1-integrin function, we stably expressed ST6Gal-I in a colon epithelial cell line lacking endogenous ST6Gal-I. ST6Gal-I expressors (with alpha2-6 sialylated beta1 integrins) exhibited up-regulated attachment to collagen I and laminin and increased haptotactic migration toward collagen I, relative to parental cells (with completely unsialylated beta1 integrins). Blockade of ST6Gal-I expression with short interfering RNA reversed collagen binding back to the level of ST6Gal-I nonexpressors, confirming that alpha2-6 sialylation regulates beta1 integrin function. Finally, we show that beta1 integrins from ST6Gal-I expressors have increased association with talin, a marker for integrin activation. Collectively, these findings suggest that beta1 hypersialylation may augment colon tumor progression by altering cell preference for certain extracellular matrix milieus, as well as by stimulating cell migration.

Adenocarcinoma↗

PubMed2002

Production of soluble integrin alpha2beta1 heterodimer complex functionally active in vitro and in vivo.

Integrin alpha2beta1, which is a membrane protein consisting of noncovalently bound alpha2 and beta1 chains, mediates cell binding to collagen and plays a role in platelet functions. DNAs encoding the chimeric proteins in which the extracellular domains of each alpha2 and beta1 chain was fused to hinge and Fc regions of human IgG(1)gamma chain were cotransfected into CHO cells. Soluble integrin alpha2beta1 (salpha2beta1) in which alpha2 and beta1 chains were covalently bound by disulfide bonds was recovered from the culture supernatant. salpha2beta1 maintained functional characteristics of cell surface alpha2beta1 as indicated by cation-dependent binding to collagen and conformational changes induced by cations or ligand. Intravenously administered salpha2beta1 in rats colocalized with collagen in inflamed microvessels. Moreover, salpha2beta1-conjugated liposome administered intravenously reduced bleeding time of the thrombocytopenic mice. These results indicated that salpha2beta1 has pharmaceutical utilities as an agent for detecting injured vessels and a component of platelet substitute.

Animals↗

PubMed2006

Wound healing in the alpha2beta1 integrin-deficient mouse: altered keratinocyte biology and dysregulated matrix metalloproteinase expression.

The alpha2beta1 integrin, a collagen/laminin receptor, is expressed at high level in the basal cell layer of the epidermis. To define the role of the alpha2beta1 integrin in wound healing, wound repair was extensively evaluated in wild-type and alpha2-null mice in vivo. In addition, the impact of alpha2beta1 integrin-deficiency on the function of primary murine keratinocytes in vitro was analyzed. Our in vivo findings demonstrate that genetic deletion of the alpha2beta1 integrin does not significantly alter the rate of re-epithelialization, collagen deposition, or tensile strength during wound closure in mice. In marked contrast to the observed similarities in wound healing, deletion of the alpha2beta1 integrin resulted in a dramatic increase in neoangiogenesis in the wound microenvironment. In contrast to in vivo studies, primary keratinocytes from alpha2-null mice adhered poorly and displayed impaired migration on type I collagen in vitro. We demonstrate that alpha2beta1 integrin-ligation negatively regulates expression of genes including matrix metalloproteinases both in vivo and in vitro. Furthermore, the changes in gene expression could potentially account for relatively normal wound healing in the alpha2-deficient mouse and our recent observation that suggests an antiangiogenic role for the alpha2beta1 integrin in vivo.

Animals↗

PubMed1999

Real-time analysis of integrin-mediated chemotactic migration of T lymphocytes within 3-D extracellular matrix-like gels.

We have developed a novel 3-D gel reconstituted with major extracellular matrix (ECM) glycoproteins to follow the dynamics of migration of human T cells locomoting, in real-time, on gradients formed by representative chemoattractants: the C-C chemokine RANTES, and the pro-inflammatory cytokine IL-2. In the absence of chemoattractants, none of the T cells migrated directionally and the levels of random migration or cell polarization were low. However, major fractions of T cells placed in IL-2 and RANTES gradients in the gels polarized immediately after exposure to the chemoattractants. Shortly after polarization, 25% of the T cells migrated, in either a random or directional fashion, towards the sources of the chemoattractants; additional 5-10% of the cells remained polarized but stationary. The number of T cells migrating directionally towards RANTES or IL-2 peaked along with the formation of the chemotactic gradients. The directional migration of T cells was increased by a short pre-exposure to low doses of IL-2, which did not alter the level of expression of the beta1 integrins. The directional migration of T cells towards IL-2 and RANTES was mediated by IL-2R and pertussis toxin-sensitive receptors, respectively, and the directional, and to a lesser degree, the random locomotion of T cells induced by both chemoattractants required intact tyrosine kinase signaling and activities of the alpha4, alpha5, and, to a lesser degree, the alpha2 and alpha6 members the beta1 integrins. Our system enables the real-time tracking of individual locomoting lymphocytes and the analysis of their dynamic interactions with ECM components and cytokines.

Cell Movement↗

PubMed1997

Altered expression of the alpha7beta1 integrin in human and murine muscular dystrophies.

The alpha7beta1 integrin is the primary laminin receptor on skeletal myoblasts and adult myofibers. It has distinct functions during muscle development and contributes to muscle structural integrity. We have studied this integrin in cases where expression of dystrophin or laminin are compromised. Immunofluorescence demonstrates an increase in alpha7beta1 in patients with Duchenne muscular dystrophy and in mdx mice that lack dystrophin. Analysis of RNA from mdx mice and from patients with Duchenne and Becker muscular dystrophies indicates that the increase in the alpha7beta1 integrin is regulated at the level of alpha7 gene transcription. In contrast, the levels of alpha7beta1 integrin are severely diminished in patients with laminin alpha2 chain congenital dystrophy muscular dystrophy and in dy/dy mice that also do not make the alpha2 laminin chain. Analysis of RNA from the hindlimbs of dy/dy mice demonstrated that in the absence of laminin alpha7 gene transcription is inhibited and limited to specific alternatively spliced isoforms. We suggest that the increased expression of alpha7beta1 integrin in the absence of dystrophin compensates for the reduced dystrophin-mediated linkage of fibers with the basal lamina and modulates the development of pathology associated with these diseases. The decrease in alpha7beta1 integrin and its transcripts in the absence of laminin likely contributes to the severe myopathy that results from laminin alpha2 chain deficiency and suggests that laminin-2 regulates expression of the alpha7 integrin gene. The role of the alpha7beta1 integrin in muscle integrity also suggests that compromised expression of this receptor may underlie as yet undefined myopathies.

Adult↗

PubMed2004

beta1-Integrins mediate enhancement of airway smooth muscle proliferation by collagen and fibronectin.

Airway smooth muscle (ASM) accumulation and enrichment of the extracellular matrix (ECM) with type I collagen and fibronectin are major pathologic features of airway remodeling in asthma. These ECM components confer enhanced ASM proliferation in vitro, but a requirement for specific integrin ECM receptors has not been examined. Here, we examined the mitogen platelet-derived growth factor (PDGF)-BB on beta1-integrin expression on human ASM cells cultured on these ECM substrates and defined the involvement of specific integrins in cell attachment and proliferation using integrin-neutralizing antibodies. PDGF-BB-dependent proliferation was enhanced two- to threefold by monomeric type I collagen or fibronectin and to a lesser extent by vitronectin; other interstitial ECM components (fibrillar type I and III collagen and tenascin-C) had no effect. Except for increased alpha3 expression induced by PDGF-BB and monomeric type I collagen or fibronectin, alpha1, alpha2, alpha4, alpha5, alphav, and alphavbeta3 integrins were unchanged compared with unstimulated cells on plastic. Blocking antibodies revealed alpha2beta1- and alphavbeta3-mediated attachment to monomeric type I collagen, whereas attachment to fibronectin required alpha5beta1. In contrast, enhancement of PDGF-BB-dependent proliferation by either monomeric type I collagen or fibronectin required alpha2beta1, alpha4beta1, and alpha5beta1 integrins. These data suggest multiple beta1-integrins regulate enhanced ASM proliferative responses.

Adult↗

PubMed2004

Molecular regulation of osteoblasts for tissue engineered bone repair.

The use of biodegradable polymers in medicine and biomedical research is increasing. A key growth area has been the use of these materials in tissue engineering, especially for guided regeneration of bone and cartilage. Our interest has been in determining the mechanisms by which cellular attachment and growth occurs on these materials. In the current study, we examined human osteoblast cell adhesion, growth, and morphologic changes on polymeric scaffolds composed of polylactic-co-glycolic acid and polylactic acid materials. We examined these characteristics in association with measurements of levels of key adhesion integrin receptors in the presence and absence of antibodies against alpha2, alpha3, alpha4, alpha5, alpha6, and beta1 subunits, and the adhesion ligand peptides RGD (Arg-Gly-Asp) and RGE (Arg-Gly-Ser). At 2 hours, results showed initial cell adhesion was considerably decreased on polylactic-co-glycolic acid and polylactic acid in the presence of the alpha2 and beta1, antibodies with a 70% adhesion rate difference observed among the groups evaluated. Higher levels of inhibition were observed on polylactic-co-glycolic acid relative to polylactic acid, which may be correlated to a higher number of cells being able to interact with the surface initially. The presence of known competitive peptide (RGD) at 2 hours, revealed its ability to block cellular adhesion to these matrices relative to the control and noncompetitive peptide RGE on polylactic-co-glycolic acid matrices. Overall adhesion rate was affected by the presence of the integrin antibodies to the alpha2, alpha3, alpha4, alpha5, alpha6, and beta1 subunits with highest differences among polylactic-co-glycolic acid relative to its control, therefore suggesting that initial osteoblastic cell adhesion to commonly used biomaterials is regulated through integrin binding.

Biocompatible Materials↗

PubMed2004

Iris pigment epithelium attachment to aged submacular human Bruch's membrane.

PURPOSE: To determine whether iris pigment epithelium (IPE) cells can attach to aged submacular human Bruch's membrane and to assess whether IPE cells express the integrin subunits that may be necessary to bind to the known extracellular matrix ligands present in Bruch's membrane. METHODS: IPE cells were seeded onto the RPE basement membrane (RPEbm) or inner collagenous layer (ICL) of aged submacular Bruch's membrane as microaggregates or were expanded in culture until enough cells could be obtained for seeding. Cell morphology and the percentage of cell coverage were determined 1 or 7 days after seeding. Messenger RNA was extracted from cultured and uncultured IPE cells and analyzed by RT-PCR. The expression of integrin subunits alpha1 to alpha6 and beta1 mRNA was examined. RESULTS: Coverage by uncultured IPE was low on both surfaces at day-1 (RPEbm, 7.9% +/- 4.8%; ICL, 5.0% +/- 2.5%) with few intact cells present. Culturing IPE improved attachment with similar coverage on both surfaces and no significant difference between day-1 (RPEbm, 89.9% +/- 9.1%; ICL, 63.4% +/- 26.5%) and day-7 (RPEbm, 97.8% +/- 2.3%; ICL, 94.7% +/- 6.6%). By day-7, cell morphology and coverage on both surfaces was variable, ranging from few intact cells to a high degree of coverage by flattened cells. All integrin subunits studied were expressed in cultured cells, whereas alpha2, alpha3, and alpha4 showed less or no expression in uncultured cells. CONCLUSIONS: Upregulation of integrin mRNA expression may be one explanation for the difference in coverage by cultured versus uncultured IPE cells. The presence of dead, dying, or flattened cells at day 7 indicates that IPE may not survive or differentiate on aged submacular Bruch's membrane.

Aged↗

PubMed2001

Integrin and FAK-mediated MAPK activation is required for cyclic strain mitogenic effects in Caco-2 cells.

Rhythmic strain stimulates Caco-2 proliferation. We asked whether mitogen-activated protein kinase (MAPK) activation mediates strain mitogenicity and characterized upstream signals regulating MAPK. Caco-2 cells were subjected to strain on collagen I-precoated membranes or antibodies to integrin subunits. Twenty-four hours of cyclic strain increased cell numbers compared with static conditions. MAPK-extracellular signal-regulated kinase (ERK) kinase inhibition (20 microM PD-98059) blocked strain mitogenicity. p38 Inhibition (10 microM SB-202190) did not. Strain rapidly and time-dependently activated focal adhesion kinase (FAK), paxillin, ERK1 and 2, and p38 on collagen. c-Jun NH(2)-terminal kinase (JNK)1 and 2 exhibited delayed activation. Similar activation occurred when Caco-2 cells were subjected to strain on a substrate of functional antibody to the alpha2-, alpha3-, alpha6-, or beta1-integrin subunits but not on a substrate of functional antibody to the alpha5-subunit. FAK inhibition by FAK397 transfection blocked ERK2 and JNK1 activation by in vitro kinase assays, but pharmacological protein kinase C inhibition did not block ERK1 or 2 activation by strain. Strain-induced ERK signals mediate strain's mitogenic effects and may require integrins and FAK activation.

Caco-2 Cells↗

PubMed1998

Isolation and characterization of laminin-10/11 secreted by human lung carcinoma cells. laminin-10/11 mediates cell adhesion through integrin alpha3 beta1.

A panel of human tumor cell lines was screened for selective expression of laminin alpha5 chain, a newly identified laminin subunit comprising laminin-10 (alpha5 beta1 gamma1) and -11 (alpha5 beta2 gamma1). The lung adenocarcinoma cell line A549 was found to express the alpha5 chain at relatively high levels but no detectable amounts of other alpha chains. The laminin variants containing alpha5 chain were purified from the conditioned medium of A549 cells by immunoaffinity chromatography using the anti-laminin monoclonal antibody 4C7 which was shown recently to recognize the laminin alpha5 chain (Tiger, C.-F., Champliaud, M.-F., Pedrosa-Domellof, F., Thornell, L.-E., Ekblom, P., and Gullberg, D. (1997) J. Biol. Chem. 272, 28590-28595). The purified laminin variants consisted of three chains with molecular masses of 350, 220, and 210 kDa. The 350-kDa chain was specifically recognized by another anti-alpha5 chain monoclonal antibody capable of recognizing denatured alpha5 chain on immunoblots, whereas the 210-kDa chain was recognized by an anti-gamma1 chain antibody. The purified alpha5 chain-containing laminin variants (hereafter referred to as laminin-10/11) were highly active in mediating adhesion of A549 cells to the substratum with potency as high as that of laminin-5 and significantly higher than those of laminin-1, laminin-2/4, or fibronectin. Adhesion to substrata coated with laminin-10/11 was specifically inhibited by anti-integrin antibodies directed against the integrin alpha3 or beta1 subunit but not by those against alpha2 or alpha6 subunit, indicating that laminin-10/11 is specifically recognized by integrin alpha3 beta1. Given the wide distribution of laminin-10/11 in the basement membrane of various tissue types and dominant expression of integrin alpha3 beta1 in most epithelial cells, specific interaction of laminin-10/11 with integrin alpha3 beta1 may play an important role in in vivo regulation of proliferation and differentiation of epithelial cells through the basement membrane.

Cell Adhesion↗

PubMed1999

Expression of beta1 and beta4 integrins in normal arachnoid membrane and meningiomas.

BACKGROUND: The aim of this work was to study the expression of alpha, beta1, and beta4 integrin subunits in meningiomas. METHODS: Seventeen atypical or anaplastic meningiomas were retrieved from the files of Hôpital de Bellevue, Saint-Etienne, France. They were compared with 17 benign meningiomas consecutively examined in 1997 and 6 schwannomas. The tumors were classified according to standard histologic criteria. Frozen sections were immunostained for alpha1, alpha2, alpha3, alpha4, alpha5, alpha6, beta1, and beta4 integrin subunits; collagen; laminin and fibronectin; cytokeratin; vimentin; neural cell adhesion molecule (NCAM); and MIB-1. RESULTS: The study included 7 fibrous meningiomas, 6 transitional meningiomas, 19 syncytial meningiomas, and 2 secretory meningiomas. The expression of alpha1, alpha3, alpha5, alpha6, and beta1 was constant. The expression of alpha1 was higher in fibrous meningiomas than in syncytial meningiomas. Only in transitional, syncytial, and secretory meningiomas was the expression of alpha2 detected. The expression of alpha2 and beta4 was associated with the expression of cytokeratin in the glandular structures of secretory meningiomas, whereas it was associated with NCAM expression in the whorls of meningothelial meningiomas. The expression of integrin receptors by tumor cells was strongly correlated with that of their respective ligands in the extracellular matrix. In invasive meningiomas, the expression of alpha3 and alpha6 by tumor cells was significantly lower. The higher the MIB-1 proliferation index, the lower the expression of alpha3. The 6 schwannomas expressed only alpha2, alpha3, alpha6, beta1, and beta4 integrins. CONCLUSIONS: Each histologic subtype of meningioma has a specific spectrum of integrin expression. The study of alpha3 and alpha6 may have prognostic value in the assessment of meningiomas. The study of the integrin profile is valuable for the differential diagnosis of fibrous meningiomas and schwannomas.

Antigens, Nuclear↗

PubMed1998

Mac-2 binding protein is a cell-adhesive protein of the extracellular matrix which self-assembles into ring-like structures and binds beta1 integrins, collagens and fibronectin.

Human Mac-2 binding protein (M2BP) was prepared in recombinant form from the culture medium of 293 kidney cells and consisted of a 92 kDa subunit. The protein was obtained in a native state as indicated by CD spectroscopy, demonstrating alpha-helical and beta-type structure, and by protease resistance and immunological analysis. It was highly modified by N- and O-glycosylation but not by glycosaminoglycans. Ultracentrifugation showed non-covalent association into oligomers with molar masses of 1000-1500 kDa. Electron microscopy showed ring-like shapes with diameters of 30-40 nm. M2BP bound in solid-phase assays to collagens IV, V and VI, fibronectin and nidogen, but not to fibrillar collagens I and III or other basement membrane proteins. The protein also mediated adhesion of cell lines at comparable strength with laminin. Adhesion to M2BP was inhibited by antibodies to integrin beta1 subunits but not to alpha2 and alpha6 subunits, RGD peptide or lactose. This distinguishes cell adhesion of M2BP from that of laminin and excludes involvement of lactose-binding galectin-3. Immunological assays demonstrated variable secretion by cultured human cells of M2BP, which was detected in the extracellular matrix of several mouse tissues.

Animals↗

PubMed2000

Potential role of alphav and beta1 integrins as oocyte adhesion molecules during fertilization in pigs.

Integrin molecules are cell adhesion molecules that are thought to be involved in sperm-oocyte interaction in rodents and humans. The objective of this study was to evaluate whether integrin molecules were present on the surface of pig oocytes, consistent with involvement in sperm-oocyte interaction in this species. Immunocytochemistry and confocal microscopy were used to evaluate the presence of beta1, and alpha1, alpha2, alpha3, alpha4, alpha5, alpha6 and alphav integrin subunits on the plasma membrane of pig oocytes. The beta1 and alphav integrin subunits were present consistently at the surface of pig oocytes; however, the remaining alpha integrin subunits evaluated were not routinely detected. The antibodies to the beta1 and alphav integrin subunits recognized appropriately sized protein bands on western blots of partially purified oocyte plasma membrane. These two antibodies also recognized oocyte plasma membrane protein isolated from a sperm plasma membrane affinity column. Sperm plasma membrane proteins of 137 and 93 kDa appeared to be the ligands for the beta1 integrin subunit as revealed by a western sandwich blot. Antibody to an extracellular domain of the beta1 integrin subunit reduced pig sperm-oocyte binding (P < 0.05), also indicating an assisting role for a beta1 oocyte integrin subunit in sperm-oocyte interaction in pigs. These results are consistent with an alphavbeta1 pig oocyte integrin interacting with a ligand on the sperm plasma membrane during fertilization.

Animals↗