Detection of invisible biological traces in relation to the physicochemical properties of substrates surfaces in forensic casework.

Touch DNA, which can be found at crime scenes, consists of invisible biological traces deposited through a person's skin's contact with an object or another person. Many factors influence touch DNA transfer, including the "destination" substrate's surface. The latter's physicochemical characteristics (wettability, roughness, surface energy, etc.) will impact touch DNA deposition and persistence on a substrate. We selected a representative panel of substrates from objects found at crime scenes (glass, polystyrene, tiles, raw wood, etc.) to investigate the impact of these characteristics on touch DNA deposition and detection. These were shown to impact cell deposition, morphology, retention, and subsequent touch DNA genetic analysis. Interestingly, cell-derived fragments found within keratinocyte cells and fingermarks using in vitro touch DNA models could be successfully detected whichever the substrates' physicochemistry by targeting cellular proteins and carbohydrates for two months, indoors and outdoors. However, swabbing and genetic analyses of such mock traces from different substrates produced informative profiles mainly for substrates with the highest surface free energy and therefore the most hydrophilic. The substrates' intrinsic characteristics need to be considered to better understand both the transfer and persistence of biological traces, as well as their detection and collection, which require an appropriate methodology and sampling device to get informative genetic profiles.

Synthesis and biological evaluation of selective Pepstatin based trifluoromethylated inhibitors of Cathepsin D.

Cathepsin D (CD) is overexpressed in several types of cancer and constitutes an important biological target. Pepstatin A, a pentapeptide incorporating two non-proteinogenic statin residues, is among the most potent inhibitor of CD but lacks selectivity and suffers from poor bioavailability. Eight analogues of Pepstatin A, were synthesized, replacing residues in P3 or P1 position by non-canonical (S)- and (R)-α-Trifluoromethyl Alanine (TfmAla), (S)- and (R)-Trifluoromethionine (TFM) or non-natural d-Valine. The biological activities of those analogues were quantified on isolated CD and Pepsin by fluorescence-based assay (FRET) and cytotoxicity of the best fluorinated inhibitors was evaluated on SKOV3 ovarian cancer cell line. (R)-TFM based analog of Pepstatin A (compound 6) returned a sub-nanomolar IC50 against CD and an increased selectivity. Molecular Docking experiments could partially rationalize these results. Stabilized inhibitor 6 in the catalytic pocket of CD showed strong hydrophobic interactions of the long and flexible TFM side chain with lipophilic residues of S1 and S3 sub-pockets of the catalytic pocket. The newly synthesized inhibitors returned no cytotoxicity at IC50 concentrations on SKOV3 cancer cells, however the compounds derived from (S)-TfmAla and (R)-TFM led to modifications of cells morphologies, associated with altered organization of F-actin and extracellular Fibronectin.

Electroactive 4D Porous Scaffold Based on Conducting Polymer as a Responsive and Dynamic In Vitro Cell Culture Platform.

In vivo, cells reside in a 3D porous and dynamic microenvironment. It provides biochemical and biophysical cues that regulate cell behavior in physiological and pathological processes. In the context of fundamental cell biology research, tissue engineering, and cell-based drug screening systems, a challenge is to develop relevant in vitro models that could integrate the dynamic properties of the cell microenvironment. Taking advantage of the promising high internal phase emulsion templating, we here designed a polyHIPE scaffold with a wide interconnected porosity and functionalized its internal 3D surface with a thin layer of electroactive conducting polymer poly(3,4-ethylenedioxythiophene) (PEDOT) to turn it into a 4D electroresponsive scaffold. The resulting scaffold was cytocompatible with fibroblasts, supported cellular infiltration, and hosted cells, which display a 3D spreading morphology. It demonstrated robust actuation in ion- and protein-rich complex culture media, and its electroresponsiveness was not altered by fibroblast colonization. Thanks to customized electrochemical stimulation setups, the electromechanical response of the polyHIPE/PEDOT scaffolds was characterized in situ under a confocal microscope and showed 10% reversible volume variations. Finally, the setups were used to monitor in real time and in situ fibroblasts cultured into the polyHIPE/PEDOT scaffold during several cycles of electromechanical stimuli. Thus, we demonstrated the proof of concept of this tunable scaffold as a tool for future 4D cell culture and mechanobiology studies.

Targeting cell-derived markers to improve the detection of invisible biological traces for the purpose of genetic-based criminal identification.

At a crime scene, investigators are faced with a multitude of traces. Among them, biological traces are of primary interest for the rapid genetic-based identification of individuals. "Touch DNA" consists of invisible biological traces left by the simple contact of a person's skin with objects. To date, these traces remain undetectable with the current methods available in the field. This study proposes a proof-of-concept for the original detection of touch DNA by targeting cell-derived fragments in addition to DNA. More specifically, adhesive-structure proteins (laminin, keratin) as well as carbohydrate patterns (mannose, galactose) have been detected with keratinocyte cells derived from a skin and fingermark touch-DNA model over two months in outdoor conditions. Better still, this combinatory detection strategy is compatible with DNA profiling. This proof-of-concept work paves the way for the optimization of tools that can detect touch DNA, which remains a real challenge in helping investigators and the delivery of justice.

In Vitro Models of Ovarian Cancer: Bridging the Gap between Pathophysiology and Mechanistic Models.

Ovarian cancer (OC) is a disease of major concern with a survival rate of about 40% at five years. This is attributed to the lack of visible and reliable symptoms during the onset of the disease, which leads over 80% of patients to be diagnosed at advanced stages. This implies that metastatic activity has advanced to the peritoneal cavity. It is associated with both genetic and phenotypic heterogeneity, which considerably increase the risks of relapse and reduce the survival rate. To understand ovarian cancer pathophysiology and strengthen the ability for drug screening, further development of relevant in vitro models that recapitulate the complexity of OC microenvironment and dynamics of OC cell population is required. In this line, the recent advances of tridimensional (3D) cell culture and microfluidics have allowed the development of highly innovative models that could bridge the gap between pathophysiology and mechanistic models for clinical research. This review first describes the pathophysiology of OC before detailing the engineering strategies developed to recapitulate those main biological features.

A biomimetic model of 3D fluid extracellular macromolecular crowding microenvironment fine-tunes ovarian cancer cells dissemination phenotype.

An early fundamental step in ovarian cancer progression is the dissemination of cancer cells through liquid environments, one of them being cancer ascites accumulated in the peritoneal cavity. These biological fluids are highly crowded with a high total macromolecule concentration. This biophysical property of fluids is widely used in tissue engineering for a few decades now, yet is largely underrated in cancer biomimetic models. To unravel the role of fluids extracellular macromolecular crowding (MMC), we exposed ovarian cancer cells (OCC) to high molecular weight inert polymer solutions. High macromolecular composition of extracellular liquid presented a differential effect: i) it impeded non-adherent OCC aggregation in suspension and, decreased their adhesion; ii) it promoted adherent OCC migration by decreasing extracellular matrix deposition. Besides, there seemed to be a direct link between the extracellular MMC and intracellular processes, especially the actin cytoskeleton organization and the nucleus morphology. In conclusion, extracellular fluid MMC orients OCC dissemination phenotype. Integrating MMC seems crucial to produce more relevant mimetic 3D in vitro fluid models to study ovarian dissemination but also to screen drugs.

Evaluation of the potential of a new ribavirin analog impairing the dissemination of ovarian cancer cells.

Epithelial ovarian cancers are insidious pathologies that give a poor prognosis due to their late discovery and the increasing emergence of chemoresistance. Development of small pharmacological anticancer molecules remains a major challenge. Ribavirin, usually used in the treatment of hepatitis C virus infections and more recently few cancers, has been a suggestion. However, Ribavirin has many side-effects, suggesting that the synthesis of analogs might be more appropriate. We have investigated the effect of a Ribavirin analog, SRO-91, on cancer cell behavioral characteristics considered as some of the hallmarks of cancer. Two human ovarian adenocarcinoma cell lines (SKOV3 and IGROV1) and normal cells (mesothelial and fibroblasts) have been used to compare the effects of SRO-91 with those of Ribavirin on cell behavior underlying tumor cell dissemination. SRO-91, like Ribavirin, inhibits proliferation, migration, clonogenicity and spheroids formation of cancer cells. Unlike Ribavirin, SRO-91 is preferentially toxic to cancer compared with normal cells. An in vitro physiologically relevant model showed that SRO-91, like Ribavirin or cisplatin, inhibits cancer cell implantation onto peritoneal mesothelium. In conclusion, SRO-91 analog effects on tumor dissemination and its safety regarding non-cancerous (normal) cells are encouraging findings a promising drug for the treatment of ovarian cancer.

Fibronectin amyloid-like aggregation alters its extracellular matrix incorporation and promotes a single and sparsed cell migration.

Fibronectin (Fn) is an extracellular matrix (ECM) multifunctional glycoprotein essential for regulating cells behaviors. Within ECM, Fn is found as polymerized fibrils. Apart from fibrils, Fn could also form other kind of supramolecular assemblies such as aggregates. To gain insight into the impact of Fn aggregates on cell behavior, we generated several Fn oligomeric assemblies. These assemblies displayed various amyloid-like properties but were not cytotoxic. In presence of the more amyloid-like structured assemblies of Fn, the cell-ECM networks were altered and the cell shapes shifted toward extended mesenchymal morphologies. Additionnaly, the Fn amyloid-like aggregates promoted a single-cell and sparsed migration of SKOV3 cancer cells, which was associated with a relocalization of αv integrins from plasma membrane to perinuclear vesicles. These data pointed out that the features of supramolecular Fn assemblies could represent a higher level of fine-tuning cell phenotype, and especially migration of cancer cells.

Synergistic effects of BMP-2, BMP-6 or BMP-7 with human plasma fibronectin onto hydroxyapatite coatings: A comparative study.

Design of new osteoinductive biomaterials to reproduce an optimized physiological environment capable of recruiting stem cells and instructing their fate towards the osteoblastic lineage has become a priority in orthopaedic surgery. This work aims at evaluating the bioactivity of BMP combined with human plasma fibronectin (FN/BMP) delivered in solution or coated onto titanium-hydroxyapatite (TiHA) surfaces. Herein, we focus on the comparison of in vitro osteogenic efficacy in mouse C2C12 pre-osteoblasts of three BMP members, namely: BMP-2, BMP-6 and BMP-7. In parallel, we evaluated the molecular binding strength between each BMP with FN using the Surface Plasmon Resonance (SPR) technology. The affinity of BMPs for FN was found totally different and dependent on BMP type. Indeed, the combination of FN with BMP-2 on TiHA surfaces potentiates the burst of gene-mediated osteogenic induction, while it prolongs the osteogenic activity of BMP-6 and surprisingly annihilates the BMP-7 one. These results correlate with FN/BMP affinity for TiHA, since BMP-6>BMP-2>BMP-7. In addition, by analyzing the osteogenic activity in the peri-implant environment, we showed that osteoinductive paracrine effects were significantly decreased upon (FN/BMP-6), as opposed to (FN/BMP-2) coatings. Altogether, our results support the use of FN/BMP-6 to develop a biomimetic microenvironment capable to induce osteogenic activity under physiological conditions, with minimum paracrine signalization. STATEMENT OF SIGNIFICANCE: The originality of our paper relies on the first direct comparison of the in vitro osteogenic potential of three osteogenic BMPs (BMP-2, -6 and -7) combined with native human plasma fibronectin delivered in solution or coated by laser transfer onto titanium hydroxyapatite surfaces. We confirm that BMP association with fibronectin enhances the osteogenic activity of BMP-2, -6 and -7, but with essential discrepancies, depending on the BMP member, and in agreement with the affinity of BMPs for fibronectin. Moreover, we bring elements to explain the origin of the BMP-2 medical life-threatening side-effects by analyzing in vitro paracrine effects. Finally, this work supports the alternative use of FN/BMP-6 to induce osteogenic activity under physiological conditions, with minimum side effects.

Amyloid-like aggregates formation by blood plasma fibronectin.

Fibronectin (FN) is a multifunctional glycoprotein of the extracellular matrix (ECM) playing critical roles in physiological and pathological cell processes like adhesion, migration, growth, and differentiation. These various functions of FN are modulated by its supramolecular state. Indeed, FN can polymerize into different types of assemblies like fibrils and aggregates. However, the mechanism of polymerization and the effects of such assemblies on cell behaviors still remain to be elucidated. Here we show that upon irreversible thermal denaturation, human blood plasma fibronectin forms high molecular weight aggregates. These compact and globular aggregates show amyloid features: they are stabilized by intermolecular b-sheets, they bind Thioflavin T and they are resistant to reducing and denaturing agents. Their characterization by electrospray ionization charge detection mass spectrometry shows that two populations can be distinguished according to the mass and charge density. Despite their amyloid features and the presence of hydrophobic patches on their surface, these aggregates are not toxic for cells. However, their binding abilities to gelatin and RGD are drastically decreased compare to native FN, suggesting possible effects on ECM-cell interactions.

Lectins as probes for assessing the accessibility of N-linked glycans in relation to the conformational changes of fibronectin.

Fibronectin, a ≈ 450-kDa protein with 4-9% (w/w) glycosylation, is a key component of extracellular matrices and has a high conformational lability regarding its functions. However, the accessibility and the role of glycosylated moieties associated with the conformational changes of fibronectin are poorly understood. Using lectins as probes, we developed an approach comprising dynamic light scattering, turbidimetry measurements, and isothermal titration calorimetry to assess the accessibility of glycosylated moieties of fibronectin undergoing thermal-induced conformational changes. Among a set of 14 lectins, fibronectin mainly reacted with mannose-binding lectins, specifically concanavalin A. When temperature was raised from 25 to 50 °C, fibronectin underwent progressive unfolding, but the conformation of concanavalin A was unaffected. Dynamic light scattering, turbidimetry measurements, and isothermal titration calorimetry showed increased concanavalin A binding to fibronectin during progressive thermal-induced unfolding of the protein core. Such data suggest that mannosylated residues are progressively exposed as fibronectin unfolds. Because oligosaccharide moieties can be differently exposed to cells, and the cell's responses could be modified physiologically or pathologically, modulation of fibronectin sugar chains could be relevant to its biological functions. Thus, lectins might be useful tools to probe the glycosylation accessibility accompanying changes in protein core folding, for which a better understanding would be of value for biological and biomedical research.

Cell cycle arrest or survival signaling through αv integrins, activation of PKC and ERK1/2 lead to anoikis resistance of ovarian cancer spheroids.

Ovarian cancer is the most lethal gynecologic cancer mainly due to spheroids organization of cancer cells that disseminate within the peritoneal cavity. We have investigated the molecular mechanisms by which ovarian cancer spheroids resist anoikis, choosing as models the 2 well-characterized human ovarian cancer cell lines IGROV1 and SKOV3. These cell lines have the propensity to float as clusters, and were isolated from tumor tissue and ascites, respectively. To form spheroids, IGROV1 and SKOV3 ovarian adenocarcinoma cells were maintained under anchorage-independent culture conditions, in which both lines survive at least a week. A short apoptotic period prior to a survival signaling commitment was observed for IGROV1 cells whereas SKOV3 cells entered G0/G1 phase of the cell cycle. This difference in behavior was due to different signals. With regard to SKOV3 cells, activation of p38 and an increase in p130/Rb occurred once anchorage-independent culture was established. Analyses of the survival signaling pathway switched on by IGROV1 cells showed that activation of ERK1/2 was required to evade apoptosis, an effect partly dependent on PKC activation and αv integrins. αv-integrin expression is essential for survival through activation of ERK1/2 phosphorylation. The above data indicate that ovarian cancer cells can resist anoikis in the spheroid state by arrest in the cell cycle or through activation of αv-integrin-ERK-mediated survival signals. Such signaling might result in the selection of resistant cells within disseminating spheroids, favoring further relapse in ovarian cancers.

Ovarian cancer ascites-derived vitronectin and fibronectin: combined purification, molecular features and effects on cell response.

BACKGROUND: Intra-abdominal ascites is a complication of ovarian cancers and constitutes a permissive microenvironment for metastasis. Since fibronectin and vitronectin are key actors in ovarian cancer progression, we investigated their occurrence and molecular characteristics in various ascites fluids and the influence of these ascites-derived proteins on cell behavior. METHODS: Fibronectin and vitronectin were investigated by immunoblotting within various ascites fluids. A combined affinity-based protocol was developed to purify both proteins from the same sample. Each purified protein was characterized with regard to its molecular features (molecular mass of isoforms, tryptophan intramolecular environment, hydrodynamic radii), and its influence on cell adhesion. RESULTS: Fibronectin and vitronectin were found in all tested ascites. Several milligrams of purified proteins were obtained from ascites of varying initial volumes. Molecular mass isoforms and conformational lability of proteins differed according to the ascites of origin. When incorporated into the cancer cell environment, ascites-derived fibronectin and vitronectin supported cell adhesion and migration with various degrees of efficiency, and induced the recruitment of integrins into focal contacts. CONCLUSIONS: To our knowledge, this is the first combined purification of two extracellular matrix proteins from a single pathological sample containing a great variety of bioactive molecules. This study highlights that ascites-derived fibronectin and vitronectin exhibit different properties depending on the ascites. GENERAL SIGNIFICANCE: Investigating the relationships between the molecular properties of ascites components and ovarian cancer cell phenotype according to the ascites may be critical for a better understanding of the recurrence of this lethal disease and for further biomarker identification.

Initial formation of IGROV1 ovarian cancer multicellular aggregates involves vitronectin.

Ovarian cancer progression is frequently associated with the development of malignant ascites. Multicellular aggregates of carcinoma cells (spheroids) found within ascites are thought to be able to promote peritoneal carcinomatosis. We have previously demonstrated the involvement of the vitronectin/alphav integrin adhesive system in the dissemination of ovarian cancer cells and continue to investigate the influence of these molecules by studying their role(s) in spheroid behavior. The aim of this study was to generate ovarian cancer multicellular aggregates and to focus on the role of vitronectin and alphav integrins in their initiation. IGROV1 cancer cells cultured in the absence of adhesive substratum formed multicellular aggregates comparable to spheroids. After 21 days, a fraction of the cells within clusters remained viable and proliferated recurrently. Within the multicellular aggregates, vitronectin and alphav integrins were co-localized at intercellular sites, suggesting their involvement in cell-cell interactions. Initial formation of IGROV1 aggregates was inhibited using anti-vitronectin and anti-alphav integrin blocking antibodies or the cyclic peptide cRGDfV. Vitronectin expression persisted during cluster disaggregation on fibronectin. These results demonstrate the ability of IGROV1 cells to generate multicellular aggregates and point to a contributory role for the vitronectin/alphav integrin system in the initial step of this process. These events could represent a prerequisite for further dissemination.

Mesothelial vitronectin stimulates migration of ovarian cancer cells.

Ovarian carcinomas, the most fatal gynaecological malignancies, are associated with poor prognosis predominantly because of a high recurrence rate. Ovarian cancer cells spread widely throughout the abdominal cavity leading to peritoneal metastasis. The influence of the mesothelial microenvironment on the biological mechanisms leading to cancer cell colonization of the mesothelium is poorly understood. This study aims to investigate whether mesothelial secretions affect the migration of ovarian cancer cells and focuses on the role of the adhesive molecule Vn (vitronectin) and its integrin receptors. An in vitro co-culture model indicated that clusters of IGROV1 and SKOV3 cells adhere to MeT-5A mesothelial cells preferentially at intercellular sites, invade the mesothelial monolayer and alter the integrity of the mesothelium. In addition, mesothelial CM (cell-conditioned medium) induces migration of IGROV1 and SKOV3 cells in Boyden chambers and wound healing assays. Furthermore, blocking molecules directed against vitronectin or its alphav integrin receptor decrease mesothelial-CM-induced migration by approximately 40% and 60-70% for IGROV1 and SKOV3 ovarian cancer cells, respectively, in Boyden chamber assays. Wound healing assays that allow cell migration to be measured over 24 h periods demonstrated that blocking molecules prevent the migration of IGROV1 and SKOV3 cells. Vitronectin is present in CM MeT-5A (mesothelial conditioned medium) and in metastatic peritoneal tissue sections. The expression of vitronectin at the periphery of mesothelial cells and within ovarian cancer cell clusters suggests a potential role for this molecule during intraperitoneal implantation of ovarian cancer cells. Vitronectin could represent a target for the development of anti-adhesive strategies to impede ovarian cancer dissemination.

Bcl-XL and MCL-1 constitute pertinent targets in ovarian carcinoma and their concomitant inhibition is sufficient to induce apoptosis.

In ovarian carcinomas, recurrence and acquired chemoresistance are the first leading causes of therapeutic failure and are responsible for the poor overall survival rate. Cisplatin exposure of sensitive cells has been previously associated with a down-regulation of Bcl-X(L) expression and apoptosis, whereas recurrence was systematically observed when Bcl-X(L) expression was maintained. Bcl-X(L) down-regulation could thus constitute an interesting chemosensitizing strategy. We showed that a Bcl-X(L)targeted RNA interference strategy efficiently sensitized chemoresistant ovarian carcinoma cells to cisplatin, but some of them were still able to re-proliferate. Considering the possible cooperation between Bcl-X(L)and MCL-1, we investigated the possibility to avoid recurrence in vitro using a multi-targeted RNAi strategy directed against these two anti-apoptotic proteins. We showed that their concomitant inhibition lead to massive apoptosis in absence of cisplatin, this multi-targeted RNAi approach being much more efficient than conventional chemotherapy. We thus demonstrated that Bcl-X(L) and MCL-1 cooperate to constitute together a strong molecular "bolt", which elimination could be sufficient to allow chemoresistant ovarian carcinoma cells apoptosis. Moreover, we demonstrated that in presence of a low concentration of cisplatin, the concomitant down-regulation of Bcl-X(L) and MCL-1 allowed a complete annihilation of tumour cells population thus avoiding subsequent recurrence in vitro in cell lines highly refractory to any type of conventional chemotherapy. Therefore, Bcl-X(L) and MCL-1 targeted strategies could constitute an efficient therapeutic tool for the treatment of chemoresistant ovarian carcinoma, in association with conventional chemotherapy.

[Epithelial ovarian tumor microecology]. / La micro-écologie des tumeurs épithéliales de l'ovaire.

Epithelial ovarian cancer remains an insidious and fatal gynecological malignancy. They are associated with a poor prognosis mainly due to a late diagnosis and to acquired chemoresistance. Cancer cell environment profoundly influences tumor development. Tumor microenvironment consists of vascular component, stromal fibroblasts, inflammatory cells and extracellular matrix. The multisite development of epithelial ovarian cancer results from molecular and cellular cross-talks between cancer cells, stromal cells and their extracellular matrix environment. These interactions involve cytokines, adhesives molecules, and proteolytic systems. This review points out the importance of micro-ecology in epithelial ovarian cancer development. For this purpose the relationships between cancer cells and their encountered microenvironments are described with suggesting some potential therapeutic perspectives.

Fluorescein isothiocyanate-labeled human plasma fibronectin in extracellular matrix remodeling.

Fluorescein isothiocyanate (FITC) is a well-known probe for labeling biologically relevant proteins. However, the impact of the labeling procedure on protein structure and biological activities remains unclear. In this work, FITC-labeled human plasma fibronectin (Fn) was developed to gain insight into the dynamic relationship between cells and Fn. The similarities and differences concerning the structure and function between Fn-FITC and standard Fn were evaluated using biochemical as well as cellular approaches. By varying the FITC/Fn ratio, we demonstrated that overlabeling (>10 FITC molecules/Fn molecule) induces probe fluorescence quenching, protein aggregation, and cell growth modifications. A correct balance between reliable fluorescence for detection and no significant modifications to structure and biological function compared with standard Fn was obtained with a final ratio of 3 FITC molecules per Fn molecule (Fn-FITC3). Fn-FITC3, similar to standard Fn, is correctly recruited into the cell matrix network. Also, Fn-FITC3 is proposed to be a powerful molecular tool to investigate Fn organization and cellular behavior concomitantly.

Cell surface overexpression of alphavbeta5 integrin impedes alphavbeta3-mediated migration of the human ovarian adenocarcinoma cell line IGROV1.

Human ovarian surface epithelium and epithelial tumors express integrin alphavbeta5, which can interact with vitronectin. In addition, in vitro acquisition of cisplatin resistance by alphavbeta3-expressing IGROV1 cells is accompanied by cell-surface expression of integrin alphavbeta5. To further explore the role of alphavbeta5 in ovarian carcinoma cells, IGROV1 cells were stably transfected with a human beta5 integrin cDNA construct, and three beta5 transfectant clones were selected for the expression of alphavbeta5 integrin at their cell surface. Despite a delayed entry in the exponential phase of growth, beta5-transfectant cells kept a proliferation ability similar to that of parental cells, while their growth rate was hindered in the presence of an anti-alphavbeta5 blocking antibody. Only simultaneous blockade of alphavbeta3 and alphavbeta5 by specific antibodies impeded the adhesion to vitronectin of beta5 transfectants and of the beta5-expressing cisplatin-resistant variant IGROV1-R10, suggesting that the two heterodimers cooperated in the regulation of this process. Cell surface expression of alphavbeta5 resulted in an attenuation of alphavbeta3-mediated migration on vitronectin. Alphavbeta5 participated to migration events in the absence of exogenous growth factors only in one transfectant clone and in IGROV1-R10 cells. Finally, the response to cisplatin was not significantly modified in beta5 transfectants when compared to IGROV1 parental cells.

Adhesion of human ovarian adenocarcinoma IGROV1 cells to endothelial cells is partly mediated by the alphav integrins-vitronectin adhesive system and induces an alteration of endothelial integrity.

Several lines of evidence suggest that vascularization plays an important role in the growth, local expansion and dissemination of ovarian epithelial tumours. However, the interaction of ovarian carcinoma cells with the endothelium remains poorly understood. To investigate adhesive events underlying this process, we used an in vitro model of cocultures between the IGROV1 human ovarian adenocarcinoma cell line and human umbilical vein endothelial cells (HUVECs). IGROV1 cells were shown to adhere rapidly on the HUVECs monolayer. Adhesion was inhibited by anti-alphav integrin and anti-Vn blocking antibodies, but not by anti-beta1 integrin antibodies. Anchorage of carcinoma cells led to the rupture of endothelial integrity, as revealed by the formation of holes in the monolayer and by the disappearance of the interendothelial VE-Cadherin network. Considering the ability of ovarian carcinoma to disseminate by a haematogenous way, these in vitro events could mimic a preliminary step for carcinoma cells crossing the endothelial barrier to extravasate.