Cell-adhesion molecules, once believed to function primarily in tethering cells to extracellular ligands, are now recognized as having broader functions in cellular signalling cascades. The CD44 transmembrane glycoprotein family adds new aspects to these roles by participating in signal-transduction processes — not only by establishing specific transmembrane complexes, but also by organizing signalling cascades through association with the actin cytoskeleton. CD44 and its associated partner proteins monitor changes in the extracellular matrix that influence cell growth, survival and differentiation.

CD44: From adhesion molecules to signalling regulators

Extracellular matrix structure.

OPN is a multifunctional cytokine and adhesion protein that contains an integrin-binding RGD sequence and additional sequences that interact with CD44v6/7 or other adhesive receptors. Its expression is increased in response to early proinflammatory cytokines and to mechanical strain in bone. The function of the secreted protein may be altered by extracellular enzymes, including thrombin and kinases. The study of OPN-null mice has revealed roles for OPN in a broad range of homeostatic (bone remodeling, tissue debridement) and pathologic (cellular immunity, wound healing, cancer metastasis) processes. While these processes seem disparate, they are linked by several common themes, including enhanced expression of OPN in response to stress or tissue injury, and stimulation of cell motility and cell survival pathways via interactions of OPN with adhesive receptors.

OPN is chemotactic for various cell types, notably monocytes/macrophages, which are attracted to sites of infection and inflammation. It is essential for cell-mediated immunity and a normal Th1 cytokine response during granuloma formation. OPN serves both to attach bone cells to bone matrix and to generate intracellular signals essential for normal osteoclast motility on bone; it may mediate osteocyte recognition of bone strain. OPN activates intracellular signaling pathways and regulates gene expression as a consequence of its interactions with its various receptors. The best-characterized is the integrin-stimulated FAK-Src-Rho pathway, which alters gelsolin function and podosome formation in osteoclasts. Identification and dissection of the signal transduction pathways and their targets are complicated by the fact that OPN can engage more than one type of receptor on the cell. For this reason, it is important to ascertain which receptors are in play in any given experimental system.

There is compelling evidence that soluble OPN can in a variety of situations help cells survive an otherwise lethal insult. Remarkably, this survival signaling is mediated by receptors that are generally considered to be receptors for ECM components. We suggest that OPN delivers an antiapoptotic “ECM-like” signal via multiple ligand-receptor interactions to cells, both adherent and nonadherent.

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Osteopontin as a means to cope with environmental insults: regulation of inflammation, tissue remodeling, and cell survival

Recent research has led to a better but as yet incomplete understanding of the complex roles osteopontin plays in mammalian physiology. A soluble protein found in all body fluids, it stimulates signal transduction pathways (via integrins and CD44 variants) similar to those stimulated by components of the extracellular matrix. This appears to promote the survival of cells exposed to potentially lethal insults such as ischemia/reperfusion or physical/chemical trauma. OPN is chemotactic for many cell types including macrophages, dendritic cells, and T cells; it enhances B lymphocyte immunoglobulin production and proliferation. In inflammatory situations it stimulates both pro- and anti-inflammatory processes, which on balance can be either beneficial or harmful depending on what other inputs the cell is receiving. OPN influences cell-mediated immunity and has been shown to have Th1-cytokine functions. OPN deficiency is linked to a reduced Th1 immune response in infectious diseases, autoimmunity and delayed type hypersensitivity. OPN's role in the central nervous system and in stress responses has also emerged as an important aspect related to its cytoprotective and immune functions. Evidence suggests that either OPN or anti-OPN monoclonal antibodies (depending on the circumstances) might be clinically useful in modulating OPN function. Manipulation of plasma OPN levels may be useful in the treatment of autoimmune disease, cancer metastasis, osteoporosis and some forms of stress.

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Osteopontin: Role in immune regulation and stress responses

The glycosaminoglycan hyaluronan (HA), a major component of extracellular matrices (ECMs), and cell surface receptors of HA have been proposed to have pivotal roles in cell proliferation, migration, and invasion, which are necessary for inflammation and cancer progression. CD44 and RHAMM (receptor of HA-mediated motility) are the two main HA receptors whose biological functions in human and murine inflammations and tumor cells have been investigated comprehensively. HA was initially considered to be only an inert component of connective tissues, but is now known as a “dynamic” molecule with a constant turnover in many tissues through rapid metabolism that involves HA molecules of various sizes: high molecular weight HA (HMW HA), low molecular weight HA (LMW HA), and oligosaccharides (oHA). The intracellular signaling pathways initiated by HA interactions with CD44 and RHAMM that lead to inflammatory and tumorigenic responses are complex. Interestingly, these molecules have dual functions in inflammations and tumorigenesis. For example, the presence of CD44 is involved in initiation of arthritis, while the absence of CD44 by genetic deletion in an arthritis mouse model increases rather than decreases disease severity. Similar dual functions of CD44 exist in initiation and progression of cancer. RHAMM overexpression is most commonly linked to cancer progression, whereas loss of RHAMM is associated with malignant peripheral nerve sheath tumor growth. HA may similarly perform dual functions. An abundance of HMW HA can promote malignant cell proliferation and development of cancer, whereas antagonists to HA-CD44 signaling inhibit tumor cell growth in vitro and in vivo by interfering with HMWHA-CD44 interaction. This review describes the roles of HA interactions with CD44 and RHAMM in inflammatory responses and tumor development/progression, and how therapeutic strategies that block these key inflammatory/tumorigenic processes may be developed in rodent and human diseases

/pdf/interactions-between-hyaluronan-and-its-receptors-cd44-rhamm-53on6xg5u9.pdf

Interactions between Hyaluronan and Its Receptors (CD44, RHAMM) Regulate the Activities of Inflammation and Cancer

Osteopontin (OPN) is a secreted glycoprotein with mineral- and cell-binding properties that can regulate cell activities through integrin receptors. Previously, we identified an intracellular form of osteopontin with a perimembranous distribution in migrating fetal fibroblasts (Zohar et al., J Cell Physiol 170:88–98, 1997). Since OPN and CD44 expression are increased in migrating cells, we analyzed the relationship of these proteins with immunofluorescence and confocal microscopy. A distinct co-localization of perimembranous OPN and cell-surface CD44 was observed in fetal fibroblasts, periodontal ligament cells, activated macrophages, and metastatic breast cancer cells. The co-localization of OPN and CD44 was prominent at the leading edge of migrating fibroblasts, where OPN also co-localized with the ezrin/radixin/moesin (ERM) protein ezrin, as well as in cell processes and at attachment sites of hyaluronan-coated beads. The subcortical location of OPN in these cells was verified by cell-surface biotinylation experiments in which biotinylated CD44 and non-biotinylated OPN were isolated from complexes formed with hyaluronan-coated beads and identified with immunoblotting. That perimembranous OPN represents secreted protein internalized by endocytosis or phagocytosis appeared to be unlikely since exogenous OPN that was added to cell cultures could not be detected inside the cells. A physical association with OPN, CD44, and ERM, but not with vinculin or α-actin, was indicated by immunoadsorption and immunoblotting of cell proteins in complexes extracted from hyaluronan-coated beads. The functional significance of OPN in this complex was demonstrated using OPN−/− and CD−/− mouse fibroblasts which displayed impaired migration and a reduced attachment to hyaluronan-coated beads. These studies indicate that OPN exists as an integral component of a hyaluronan-CD44-ERM attachment complex that is involved in the migration of embryonic fibroblasts, activated macrophages, and metastatic cells. J. Cell. Physiol. 184:118–130, 2000. © 2000 Wiley-Liss, Inc.

N. Suzuki

Papers

Intracellular osteopontin is an integral component of the CD44-ERM complex involved in cell migration.