Introduction Bone marrow BM derived endothelial progenitor
Bone marrow (BM)-derived endothelial progenitor congo red (EPCs) circulate in peripheral blood and play an important role in the formation of new blood vessels (Asahara et al., 1997; Takahashi et al., 1999; Zhou et al., 2013; Li et al., 2011). This process occurs in response to a variety of growth factors, cytokines, and chemokines, which cause EPCs to migrate towards target tissues and differentiate into endothelial cells. Transplantation studies have revealed that EPCs incorporate into active neovasculature in ischemic hind limbs and myocardium, and growing tumors (Zhou et al., 2013; Otani et al., 2002; Kalka et al., 2000). Number and migratory activity of circulating EPCs have been reported to be inversely correlated with risk factors for artery diseases, but directly correlated with tumor growth and metastasis (Vasa et al., 2001; Naik et al., 2008). Therefore, EPCs have been exploited for new intervention strategies in therapeutic neovascularization (such as ischemia) and certain angiogenesis-dependent diseases (such as cancer) (Li Calzi et al., 2010).
An accumulating body of evidence indicates that the chemokine receptor CXCR2 promotes neovascularization and endothelial cell migration (Li et al., 2003; Schraufstatter et al., 2001). CXCR2 is a G protein-coupled receptor that can be activated by the ELR+ CXC chemokines, including CXCL1 (keratinocyte-derived chemokine; KC/murine CXCL1) and CXCL2 (macrophage inflammatory protein-2, MIP-2/murine CXCL2/3) (Olson and Ley, 2002). Recently, it has been reported that CXCR2 receptor and its cognate ligands mediate EPC recruitment and angiogenesis in injured endothelium (Hristov et al., 2007), ischemic myocardium (Kocher et al., 2006), and allergic airway disease (Jones et al., 2009; Imaoka et al., 2011). Also, it has been shown that mobilization of BM-derived EPCs was attenuated in CXCR2 knockout mice with pancreatic cancer (Li et al., 2011).
PDZ (PSD-95/DlgA/ZO-1) domains are modular protein interaction domains that form peptide-binding clefts and typically mediate interactions with the carboxyl termini of other proteins that terminate in consensus binding motifs (referred to as PDZ motif) (Li and Naren, 2010). A variety of PDZ domain-containing proteins (also referred to as PDZ scaffold or adaptor proteins) have been documented to nucleate the formation of compartmentalized multiprotein complexes critical for specific cell signaling (Li et al., 2005, 2007, 2010). Of note, both human and murine CXCR2 possess a consensus PDZ motif at their carboxyl termini. We have recently reported that the PDZ motif (TTL) of CXCR2 is involved in the regulation of intracellular signaling and cellular functions in neutrophils (Wu et al., 2012) and pancreatic cancer cells (Wang et al., 2013). We showed that CXCR2 and its downstream signaling molecule PLC-β are clustered by a PDZ adaptor protein NHERF1 which scaffolds a CXCR2–PLC-β complex through PDZ-mediated interactions (Wu et al., 2012; Wang et al., 2013). We also reported the crystal structure of the complex formation by X-ray crystallization, which provides the structural basis for CXCR2-mediated cellular functions in neutrophils and pancreatic cancer cells (Lu et al., 2013; Jiang et al., 2014). However, the putative role and the molecular mechanisms that underlie the functional significance of potential PDZ-based CXCR2 complexes in EPC mobilization, homing, and incorporation into neovasculature have not been determined. The objective of this study was to define the biological role and molecular mechanisms of the PDZ motif of CXCR2 in the regulation of EPC homing and angiogenesis.
Materials and methods
Discussion A large body of evidence has accumulated to indicate that EPCs, a subpopulation of vascular precursor cells in the blood, have the capacity to migrate, proliferate, and differentiate into mature endothelial cells in a variety of systems. EPCs contribute to postnatal neovascularization and play important roles in both ischemic vascular repair and tumor growth (Asahara et al., 1997; Bagley et al., 2003). The homing of EPCs to neovascular areas requires a coordinated sequence of multistep events, including chemoattraction, adhesion, and invasion, before incorporation and differentiation into endothelial cells (Vajkoczy et al., 2003). A number of locally produced factors, including cytokines, chemokines, and growth factors, are involved in orchestrating these processes. Several key angiogenic chemokines (CXCL1, CXCL2, CXCL7, CXCL8, CXCL12) and their cognate receptors (CXCR2, CXCR4) have been reported to regulate EPC mobilization, recruitment, and firm adhesion during EPC homing to the sites of arterial injury (Hristov et al., 2007; Kocher et al., 2006; Jones et al., 2009; Imaoka et al., 2011; Chen et al., 2012; Cheng et al., 2010). Although the role of EPCs in supporting postnatal neovascularization is under intensive investigation, the mechanisms and factors regulating these signaling events are still not fully understood.