P2Y12 receptor antagonists and AR receptor agonists regulates Protein Disulfide Isomerase secretion from platelets and endothelial cells.

Secretion of PDI from platelets and endothelial cells is an important step of all thrombotic events. In the absence of extracellular PDI thrombus formation and fibrin generation may be impaired. Thrombin-mediated PDI secretion is regulated by the stimulation of P2Y12 receptors. This paper provides evidences that P2Y12 antagonists or AR agonists may modulate release of PDI molecules from platelets and with less efficiency from endothelial cells. Moreover P2Y12 antagonization or AR agonization modulates platelet-endothelial interaction. We prove that combinations of P2Y12 antagonists and AR agonists inhibit platelet-dependent adhesion of cancer cells to endothelium and attenuate cancer cell invasiveness, but longer exposition to AR agonists may stimulate migration of invasive breast cancer cells through endothelium thus leading to increased metastasis.

Modifications of disulfide bonds in breast cancer cell migration and invasiveness.

Cancer metastasis involves the adhesion of cancer cells to the endothelium. This process can be mediated by integrins which are surface receptors responsible for interactions with ECM proteins. Integrins ß1 and αVß3 represent factors are involved in cancer progression and metastasis. Activation of integrins can be promoted by thiol-disulfide exchanges initiated by Protein Disulfide Isomerase (PDI). The purpose of this study was to prove the involvement of disulfide rearrangements in the molecules of integrins in the course of cancer cell adhesion and migration through the endothelium. We present the evidence which proves that highly metastatic MDA-MB-231 breast cancer cell lines adhere to endothelial cells are more effective than non-invasive MCF-10A and MCF-7 cell lines and that the attachment of MDA-MB-231 to the endothelium can be attenuated either by the agents blocking free thiol groups (DTNB, cystamine or PCMBS) or by PDI inhibitors (Q3Rut, 16F16 or PACMA-31). Furthermore, we prove that the transendothelial migration of MDA-MB-231 cells and contraction of collagen can be blocked by thiol blockers or PDI inhibitors and that these factors affect exposition of free thiols on integrin molecules.

Contribution of activated beta3 integrin in the PDI release from endothelial cells.

Protein disulfide isomerase (PDI) is an abundant reticulum endoplasmic protein but also acts as an important functional regulator of some extracellular surface proteins. Recent studies suggest that PDI plays a role in integrin activation and thrombus formation. The aim of this study was to examine whether activation of integrin is the first stage leading to release of PDI from the subcellular compartments of endothelial cells to extracellular space. Our results show that endothelial cells which adhere to fibronectin or fibrinogen release significantly more PDI than those which adhere to poly-L-lysine. Cells treated with RGD peptide, Src and FAK kinase inhibitors and anti alphaVbeta3 antibody display lower PDI secretion. The destruction of the actin cytoskeleton of endothelial cells by cytochalasin D inhibits PDI release. When the endothelial cells adhere to fibrinogen or fibronectin, PDI and alphaVbeta3 gain free thiol groups. Our data suggest that upon activation of integrins, PDI is released from endothelial cells and forms a disulfide bond complex with alphaVbeta3 integrin.

The role of Protein Disulfide Isomerase and thiol bonds modifications in activation of integrin subunit alpha11.

Integrins belong to a family of transmembrane receptors that mediate cell migration and adhesion to ECM. Extracellular domains of integrin heterodimers contain cysteine-rich regions, which are potential sites of thiol-disulfide exchanges. Rearrangements of extracellular disulfide bonds regulate activation of integrin receptors by promoting transition from an inactive state into a ligand-binding competent state. Modifications of integrin disulfide bonds dependent on oxidation-reduction can be mediated by Protein Disulfide Isomerse (PDI). This paper provides evidences that binding to integrin ligands initiate changes in free thiol pattern on cell surface and that thiol-disulfide exchange mediated by PDI leads to activation of integrin subunit α11. By employing co-immunoprecipitation and confocal microscopy analysis we showed that α11ß1 and PDI create complexes bounded by disulfide bonds. Using surface plasmon resonance we provide biochemical evidence that PDI can interact directly with integrin subunit α11.

[Role of protein disulfide isomerase in activation of integrins]. / Rola izomerazy disiarczkowej w aktywacji integryn.

Integrins belong to a large family of transmembrane cell adhesion receptors that communicate biochemical and mechanical signals in a bidirectional manner across the plasma membrane. Integrins and their ligands play a crucial role in a number of physiological and pathological processes, including cell migration, cell differentiation, hemostasis, adhesion, angiogenesis, cancer, cell invasiveness and wound healing. Intracellular signals switch integrins into a ligand-competent state as a result of conformational changes within the integrin molecule. Binding of extracellular ligands induces structural changes that can transmit signals to the cell interior. Transition of integrins from an inactive to a ligand binding state involves rearrangement of the disulfide bonding pattern. The rearrangement of disulfide bonds is modulated by protein disulfide isomerase (PDI). PDI has been found on the surface of several types of cells, including endothelial cells, hepatocytes, cancer cells, pancreatic cells and B cells. PDI was identified on the platelet surface, where it plays an important role in platelet reactions such as adhesion, aggregation and secretion. PDI was found to directly interact with integrins. Disulfide-thiol exchange mediated by PDI appears to be involved in the conformational changes in integrin activation. In this report we describe the structure of integrin and the role of disulfide bond rearrangement in its activation.