Alarmin-induced cell migration

Abstract

Alarmins are endogenous, constitutively available, damage-associated molecular patterns that upon release can mobilize and activate various leukocytes for the induction of innate and adaptive immune responses. For our immune system to function appropriately, it relies on navigating various leukocytes to distinct places at the right time. The direction of cell migration is determined by chemotactic factors that include classical chemoattractants, chemokines, certain growth factors, and alarmins. This viewpoint provides an overview of alarmin-induced cell migration. Alarmins are capable of inducing the migration of diverse types of leukocytes and nonleukocytes either directly by triggering specific receptors or indirectly by inducing production of chemokines through the activation of various leukocytes via pattern recognition receptors. The receptors used by alarmins to directly induce cell migration can either be Gαi protein-coupled receptors or receptors such as the receptor for advanced glycation end products; however, the intracellular signaling events responsible for the direct chemotactic activities of alarmins are, to date, only partially elucidated. Given that alarmins act in concert with chemokines to regulate the recruitment and trafficking of leukocytes, these damage-associated molecular patterns are potentially involved in diverse biological processes as discussed in this viewpoint.

Introduction

The migration of single cells is important for many physiologic processes such as ontogenic development, organogenesis, hematopoiesis, tissue regeneration, and immune responses. Cell migration is also involved in pathological conditions such as autoimmune disorders, vascular disease, and tumor metastasis. The ability of the immune system to respond appropriately to microbial invasion, tissue damage, and other insults relies to a large extent on the mobilization/recruitment of various leukocytes and progenitor cells to the right place at the right time [1,2]. Migration of cells in vivo is controlled by many sequential interactions involving adhesion molecules, glycosaminoglycans, chemotactic factors, and their receptors [1–5].

For cells to move directionally, they must first acquire a polarized morphology where F-actin is primarily enriched at the front and myosin II is assembled on the sides and at the back of the cell [6]. Subsequently, the polarized cells undergo a highly coordinated cycle of protrusions and retractions that are coupled with traction provided by the formation and release of adhesive contacts with the extracellular matrices [6]. Cells must be able to determine where and when protrusions, retractions, and adhesions have to occur to migrate to the correct location, which is established by chemotactic gradients. Chemotactic factors are comprised of classical chemoattractants such as formyl peptides and anaphylatoxins (e.g. C5a), chemokines including CXC, CC, CX3C, and XC chemokines, and growth factors such as EGF and vascular endothelial growth factor [1, 4, 7, 8]. Classical chemoattractants and chemokines provide extracellular cues by signaling predominantly through Gαi protein–coupled receptors (GiPCRs), while growth factors do so by signaling through their corresponding receptors [4,7,8].

A more recently identified type of chemotactic factor is the alarmin family. Alarmins are structurally distinct endogenous mediators that, upon release and gaining access to immune cells, can activate the immune system by inducing the recruitment and activation of various leukocytes, particularly APCs, including DCs [9–11]. Consequently, alarmins are capable of inducing both innate and antigen-specific host immune responses. Most alarmins are constitutively expressed and stored in intracellular compartments such as the nucleus, cytoplasm, or granules. The expression of some alarmins can also be upregulated by microbial products, cytokines, and stress. During microbial infection and/or tissue injury, alarmins rapidly become available extracellularly as a result of degranulation, passive release due to cell necrosis, or active release in response to inducing agents [9–11].

Known alarmins are multifunctional and can be classified into eight distinct molecular categories (Table 1), including defensins (e.g. α- and β-defensins), cathelicidin (e.g. human LL-37 or mouse cathelicidin-related antimicrobial peptide (CRAMP)), eosinophil-associated ribonucleases (e.g. eosinophil-derived neurotoxin (EDN)), nuclear-binding proteins (e.g. high-mobility group box-1 protein (HMGB1), high-mobility group nucleosome-binding protein 1 (HMGN1), and the cytokines IL-1α and IL-33), HSPs (e.g. HSP60, HSP70), saposin-like granulysin, ion-binding proteins (e.g. lactoferrin, S100 proteins), and nucleotides/metabolites (e.g. ATP, uric acid). Several alarmins, including HMGB1, S100A8/9, ATP, and uric acid, were found to not only play a chemotactic role but to also function as damage-associated molecular patterns (DAMPs) since they are released as a result of cell injury/death and can perpetuate immune responses [10–12]. The term DAMP was proposed in 2004 to designate hyppos (biological molecules with hydrophobic portions) capable of initiating repair, remodeling, and immune responses [12]. DAMPs are broadly defined and include both endogenous molecules engaged in host defense and metabolism (HSPs, uric acid, etc.) and exogenous pathogen-associated molecular patterns such as LPS, flagellin, and bacterial DNA [12].

Table 1.

Target cells and receptors of alarmin-induced cell migration.

Alarmin		Target cell	Receptor
Category	Member		GiPCR	Non-GiPCR
Defensin	α-Defensin family	MC [21], Mo/Mφ [16, 21], DC [17], T [15, 17, 21]	n.d.	n.d.
	β-Defensin family	MC [22], Mo/Mφ [26, 27], DC [18, 19, 23, 28], T [18], EPC [37, 70], Ep [75]	CCR6 [18–20, 23, 70], CCR2 [26, 27]	EGFR [75]
Cathelicidin	LL-37/CRAMP	PMN [32], MC [33, 34], Mo [32], DC [38], T [32], MSC [35, 36] Ep [35, 37]	FPRL1/FPR2 [32, 35, 36, 38], MrgX2 [34]	EGFR [37]
EAR	EDN/EAR2	DC [40]	n.d.	n.d.
Nuclear-binding protein	HMGB1	PMN [50], Mφ [51], DC [52], MSC [53–55], EPC [56], SMC [57, 58], Ep [59, 60]	CXCR4 [89, 99]	RAGE [50–53, 56, 72, 73]
	HMGN1	Mo, DC [49]	n.d. [59]	n.d.
	IL-33,IL-lα	PMN [64], T [63]	n.d.	ST2 [63]
HSP	HSP60, 70	DC, NK [41]	n.d.	n.d.
Saposin-like	Granulysin	Mo [39], DC [42], T [39]	n.d.	n.d.
Ion-binding protein	S100a7,8,12,15	PMN [43, 47], MC [44], Mo/Mφ [46, 47]	n.d.	RAGE [46]
	Lactoferrin	Mo/Mφ [45]	n.d.	n.d.
Nucleotide/metabolite	ATP	PMN, Eo, Mo/Mφ, DC, EC, SMC [66]	P2Y2, P2Y6, P2Y12 [66]	P2X7 [66]
	Uric acid	Eo [65]	n.d.	n.d.

CCR: CC chemokine receptor; EAR: eosinophil-associated ribonuclease; EC: endothelial cell; Ep: epithelial cell; FPRL1: formyl peptide receptor-like 1 receptor; Mφ: macrophage; Mo: monocyte; MrgX2: Mas-related gene X2; MSC: mesenchymal stem/stromal cell; PMN: polymorphonuclear neutrophil; T, T cell. n.d. = not determined.

Given the dual roles of defensins, cathelicidins, EDN, and HMGB1, the term alarmin was coined, also in 2004, to classify a set of endogenous mediators that possess the dual capacities of promoting host defenses against dangers by inducing the migration/recruitment and activation of APCs and consequently are capable of initiating/enhancing innate and adaptive host immune responses [9, 13]. Alarmins, like cytokines, can be dangerous if produced in excess under inappropriate circumstances such as during severe injury, autoimmunity, and tumor progression. Since alarmins and DAMPs overlap in terms of release and immunostimula-tory effect, alarmins can be considered an endogenous subset of DAMPs [10, 14]. It has also become clear since the initial classification that alarmins are involved in the induction of cell migration, in vivo recruitment, and cell activation through multiple mechanisms.

Alamins directly induce cell migration

The chemotactic effects of alarmins

Alarmins are chemotactic for diverse types of leukocytes as well as nonleukocytes (Table 1). Both human and mouse α-and β-defensins are chemotactic for immature DCs, monocytes/macrophages, mast cells (MCs), and certain subsets of T lymphocytes [15–27]. The leukocyte chemotactic activity of defensins appears to be universal across vertebrate species since defensins from bovine and fish species are also chemotactic [28,29]. Cathelicidins can chemoattract many subsets of leukocytes as well as nonleukocytes such as mesenchymal stromal cells and keratinocytes [30–38]. EDN, HSPs, granulysin, S100 proteins, and lactoferrin are chemotactic for various subsets of leukocytes [39–48]. The recently identified alarmin HMGN1 is important for the recruitment of DCs in vivo [49] and possesses direct chemotactic activity for monocytes and DCs (D. Yang et al., unpublished results). HMGB1, which belongs to the nuclear-binding protein category as does HMGN1, is a multi-functional alarmin that has been shown to induce migration of the widest spectrum of target cells, including neutrophils [50], monocytes/macrophages [51], DCs [52], mesoangioblasts [53], mesenchymal stromal cells [54, 55], endothelial progenitor cells (EPCs) [56], smooth muscle cells (SMCs) [57, 58], fibroblasts and keratinocytes [59], and certain tumor cells [60]. HMGB1 also promotes the outgrowth of neurites and the motility of neurons [61, 62]. IL-33, a nuclear-binding protein alarmin, is reported to be chemotactic for Th2 T lymphocytes and neutrophils [63, 64]. Nucleotides and their metabolites, including uric acid, are the only type of alarmins that are not protein in nature. Uric acid has thus far only been reported to induce chemotaxis of eosinophils [65], while ATP is chemotatic for many leukocytes and nonleukocytes (reviewed recently in[66]).

The chemotactic effects of alarmins on various target cells are often demonstrated in vitro by a Boyden chamber-based multi-wall chemotaxis assay, a reliable method widely used for investigating the migration of cells in response to many chemotactic factors including chemokines. The capacity of various alarmins to induce in vivo cell recruitment can be demonstrated by injecting alarmin(s) into air-pouch, peritoneal cavity, or solid tissue and subsequently quantifying the type and number of leukocytes attracted into the injection sites. For example, the T-cell attracting capacity of human neutrophil-derived α-defensin was confirmed by the accumulation of human CD3-positive T cells at the subcutaneous site 4 h after injection of α-defensin in an experimental model system [15]. The in vivo capacity of HMGB1 to induce the recruitment of mesoangioblasts was determined by the migration of intraartery injected mesoangioblasts toward HMGB1-loaded heparin-Sepharose beads implanted in the muscle of experimental mice [53]. The capacity of EDN or cathelicidin to chemoattract APCs in vivo was demonstrated by the recruitment of DCs and macrophages into air-pouches after intrapouch administration of the respective alarmins [40,67]. The most often used in vivo model is the peritonitis model, which has been used to verify the in vivo leukocyte-recruiting capacity of lactoferrin, granulysin, HMGB1, and HMGN1 [42,45,49,68].

What appears to be common among most alarmins is the capacity to induce the migration of APCs including monoyctes, macrophages, and DCs (Table 1). It is likely that most alarmins participate in regulating the recruitment and trafficking of DCs. Since the characterization of alarmin-induced cell migration is still incomplete, much more has to be elucidated before a complete picture can be painted with regard to the target cell spectrum of all alarmins.

Receptors that mediate the chemotactic effects of alarmins

The chemotactic cell migration induced by most alarmins can be inhibited by pre-treatment of the target cells with pertussis toxin, a bacterial toxin capable of preventing G proteins from interacting with GiPCRs on the cell membrane by catalyzing the ADP-ribosylation of the α_i subunits of the heterotrimeric G protein [17, 18, 21,22,25,32,33,35,36,39,40,45,52,67]. This indicates that the direct chemotactic effects of many alarmins are mediated by GiPCRs (Table 1). Some alarmins use more than one GiPCR, e.g. β-defensin 2 uses both CCR6 and CCR2 [18,19,26,27]. Furthermore, GiPCR usage by alarmins overlaps with that of chemokines; e.g. the β-defensin 2 and 3 share CCR6 with CCL20 [18, 19, 23, 24, 69–71], while the β-defensin 3 and 14, which were more recently reported to also use CCR2 [26, 27], share CCR2 with a number of CC chemokines such as CCL2, CCL7, CCL8, and CCL12 [1,4,7].

The capacity of human and mouse cathelicidin to induce the migration of most target cells is mediated by FPRL1 (FPR2 in mouse) [32,35,36,67] and it has recently been shown that LL-37 induces MC migration and degranulation through MrgX2, a GiPCR belonging to Mas-related gene family [34]. ATP uses several GiPCRs for inducing the migration of various target cells, such as P2Y2 on DCs, eosinophils, and fibroblasts, P2Y6 on monocytes and P2Y12 on microglia and SMCs [66]. The identity of GiPCRs responsible for mediating the chemotactic effects of α-defensins, eosinophil-associated ribonucleases, HSPs, HMGN1, granulysin, lactoferrin, and uric acid, remain to be identified (Table 1).

Receptors other than GiPCRs are also used by alarmins to induce chemotactic migration (Table 1). HMGB1-induced migration of neutrophils, monocytes, DCs, mesoangioblasts, endothelial progenitors, and glioblastoma cells is dependent on the presence of the receptor for glycation end products (RAGE) [50–53, 56, 72, 73]. While S100A8- and S100A15-induced migration of neutrophils and macrophages is mediated by GiPCRs [46–48], monocyte migration induced by S100A7 is RAGE-mediated [46]. IL-33-induced cell migration appears to be mediated by ST2, an IL-1 receptor like-1 receptor [63, 74]. The EGF receptor may mediate the cell-attracting effect of some alarmins. The migration of keratinocytes in response to LL-37 appears to be mediated by transactivation of EGF receptor [37]. The migration of keratinocytes induced by several β-defensins is also reported to involve EGF receptor transactivation in a pertussis toxin-sensitive manner [75], which suggests that an as-yet-unidentified GiPCR may also be involved. The interaction between LL-37 and insulin-like growth factor 1 receptor on MCF-7, a human breast cancer cell line, also induces cell migration, suggesting that LL-37 may also use this receptor to promote the migration of certain tumor cells[76].

Intracellular signaling pathways

Several intracellular signaling cascades driving cell migration are triggered by alarmins. Cathelicidins elevate intracellular Ca²⁺ in neutrophils, monocytes, and MCs [30, 32, 33, 67]. LL-37 induces signaling through insulin-like growth factor 1 receptor to activate ERKs, phosphatidylinositol-3 kinase (PI3K)-Akt pathways, both of which participate in the migration and motility of fibroblasts and breast cancer cells [76]. The migration of intestinal epithelial cells induced by human β-defensin 2 is accompanied by an increase of intracellular Ca²⁺, activation of RhoA, and PI3K [71]. HMGB1-induced migration of glioblastoma cells, endothelial cells, and mesoangioblasts depends, at least in part, on the activation of ERKs [73, 77, 78]. HMGB1 triggers cytoskeleton reorganization in SMCs, which is necessary for their migration [57]. On the other hand, HMGB1-stimulated migration of human chondrosarcoma cells and SMCs requires the activation of the PI3K-Akt signaling pathway [58,60]. Defensins also induce the activation of the PI3K-Akt signaling pathway in intestinal epithelial cells in the course of promoting their migration [71]. Overall, it appears that alarmins, in the process of inducing the migration of diverse types of cells, trigger the activation of PI3K/Akt, ERKs, small GTPases (e.g. Rho, Rac, etc.), PKC, and elevation of intra-cellular Ca2⁺.

For cell migration induced by alarmins, the intracellular signaling events that connect the receptor (s) and intracellular signaling messengers (e.g. PI3K/Akt, ERKs, PKC, Rho, Ca2⁺, etc.) still need to be determined. Since the predominant receptors mediating the chemotactic effects of alarmins are GiPCRs and RAGE, and the signaling pathways of chemotactic receptors (e.g. FPRL1, CCR2, CCR6) or RAGE have been elucidated in some detail [1, 4, 7, 8], it is likely that alarmins trigger similar intracellular signaling pathways to those of chemokines and AGEs for the induction of cell migration as shown in Figure 1. Thus binding of an alarmin with its GiPCR presumably results in the activation of heterotrimeric G-protein(s) that dissociate into activated Gαi and Gβγ subunits. Activated Gαi and GβGγ trigger a series of reactions, commencing with the activation of phospholipase C (PLCβ), PI3K, and small GTPases (Rac and Rho) [1, 4, 7, 8]. Ligand engagement of RAGE leads to activation of PI3K [79] and Rac [80]. PLCβ hydrolyses phosphatidylinositol-4, 5-bisphosphate (PIP2), generating inositol 1,4,5-trisphosphate (IP3), and diacylglycerol. IP3 induces the release of calcium from intracellular stores, and together with diacylglycerol, activates PKC. Activation of PI3K in turn phosphorylates Akt. Rac and Rho initiates a series of reactions that lead to the activation of ERKs, LIM (named by the initials of the three homeodomain proteins Lin11, Isl-1, and Mec-3 in which it was first discovered) domain kinase (LIMK), and myosin light chain. These protein kinases act cooperatively to rearrange cytoskeleton fibers leading to cell migration (Fig. 1).

Figure 1.

Pathways of alarmin-induced cell migration/recruitment. Alarmins can directly induce cell migration by engaging either a GiPCR or RAGE (left), or indirectly by stimulating the production of chemokines, growth factors, and adhesion molecules (right). Alarmin engagement with its GiPCR would lead to the activation and dissociation of heterotrimeric G proteins into Gαi and Gβγ subunits, which in turn, trigger the activation of PLCβ, PI3K, Rac, and Rho. PLCβ hydrolyses PIP2, generating IP3 and diacylglycerol. IP3 elevates intracellular calcium, which together with diacylglycerol activates PKC. Activation of PI3K in turn phosphorylates Akt. Rac and Rho initiates a series of reactions that lead to the activation of ERKs, LIMK, and myosin light chain. Ligand engagement of RAGE leads to activation of PI3K and Rac, and subsequent activation of Akt, ERKs, and LIMK. The kinases (Akt, ERKs, LIMK, MLC, and PKC) work cooperatively to rearrange cytoskeleton fibers, and ultimately to enable cell migration.

Alarmins indirectly promote cell migration and recruitment

Alarmins can activate many cell types, often by triggering a pattern recognition receptor such as TLRs [9–11, 14, 49, 81]. This leads to the production of chemotactic factors and adhesion molecules, both of which participate in promoting cell migration and/or recruitment [1–5,8]. For example, treatment of airway epithelial cells with human α-defensin promotes the production of CXCL8 [82]. LL-37 treatment of endothelial cells leads to the production of CCL2 [83]. Several human β-defensins can stimulate keratinocytes to produce a number of chemokines including CXCL10, CCL2, CCL20, and CCL5 [75]. HSP70 released from heat-shocked tumor cells stimulates the production of chemokines (CXCL10, CCL2, and CCL5) that induces the infiltration of DCs and T cells [84]. Activation of macrophages or DCs by EDN, HMGB1, HMGN1, granulysin, S100a8/9, and uric acid leads to the production of many chemokines such as CXCL5, CXCL7, CXCL8, CXCL9, CXCL10, CXCL12, CCL1, CCL2, CCL3, CCL5, CCL7, and CCL8 [39,42,49,52,65,85–89]. These chemokines, in turn, promote the migration and recruitment of cells positive for the corresponding chemokine receptors including CXCR1, CXCR2, CXCR3, CCR1, CCR2, CCR5, and CCR6. LL-37 is capable of inducing the generation of vascular endothelial growth factor, which, in turn, promotes angiogenesis by facilitating the recruitment of multipotent mesenchymal stromal cells [36].

LL-37-treated EPCs migrate better than nontreated progenitor cells to the injured area due to the upregulation of E-selectin and P-selectin glycoprotein ligand-1 [90]. LL-37 also enhances ICAM-1 expression by endothelial cells [83]. HMGB1 is capable of inducing α5β1 integrin expression by human chondrosarcoma cells [60]. HMGB1 not only upregulates the expression of ICAM-1, and the β1 and β2 inte-grins on EPCs, but it also increases the affinity of these integrins, contributing to the enhanced adhesion and recruitment of EPCs [56]. Human β-defensin 2 has recently been shown to induce the arrest of Th17 cells on inflamed endothelial cells in an ICAM-1-dependent manner [91]. Therefore, alarmins have the capacity to regulate the expression or activation of various adhesion molecules on both endothelial cells and target cells and thus can promote the migration of target cells.

Overall, alarmins can induce cell migration directly through interacting with their GiPCRs or RAGE, or indirectly by stimulating the production of chemokines, growth factors, and adhesion molecules often via activating the corresponding pattern recognition receptor (Fig. 1). The mechanism used by a given alarmin depends on the type of target cell involved. It is also likely that a given alarmin induces the in vivo trafficking of target cells by more than one mechanism. A good example would be LL-37-induced migration and activation of phagocytes, MCs, DCs, and epithelial cells. LL-37/CRAMP induces the migration of neutrophils and monocytes through the use of FPRL1/FPR2 [32, 67], however, its capacity to stimulate the migration of MCs is mediated by MrgX2 [34]. In contrast, LL-37 can transactivate EGFR leading to the activation of ERKs and production of IL-8[92], which would in turn promote the recruitment of neutrophils by the use of CXCR1 and CXCR2. Furthermore, LL-37 also forms complexes with DNA or RNA to promote the activation of DCs by triggering TLR7, TLR8, and TLR9, which contributes to the recruitment of inflamma-tory cells during psoriasis and skin wound healing [93, 94]. Thus, alarmins presumably promote the migration and recruitment of diverse types of cells via multiple mechanisms.

Biological implications

What is the relationship between alarmins and other chemotactic factors? For example, many alarmins can induce the migra tion of DCs (Table 1) as can many classical chemoattractants and chemokines [7, 95, 96]. Knockout of FPR2, a GiPCR for LL-37/CRAMP [32, 67], impairs the recruitment of DCs to the site of inflammation [38], suggesting a nonredundant contribution of cathelicidin to DC recruitment. Knockout of HMGN1 greatly reduces DC accumulation at the site of immunization [49]. The recruitment of DC precursors to the skin is dependent on CCR2, indicating the importance of the chemokine CCL2 in the process [97]. Both CCL20 and many defensins use CCR6 as the receptor to induce the migration of immature DCs [7, 18, 19, 70, 95, 96, 98]. Therefore, both alarmins and chemokines potentially contribute to migrational navigation of immature DCs from the blood to sites of inflammation and antigen presence. In addition, HMGB1 can form complexes with CCL12, which induces DC migration by triggering CXCR4 and RAGE [89, 99]. Mature DC homing to draining lymph nodes requires not only HMGB1-RAGE [100], but also chemokines including CCL19, CCL21, and CXCL12 [7, 95–97]. Thus, alarmins are likely to act in concert with other chemo-tactic factors to regulate the in vivo recruitment of various types of leukocytes.

The capacity of alarmins to induce cell migration and recruitment plays important roles in many biological processes. HMGB1-promoted skin wound repair depends on its capacity to chemoattract fibroblasts and keratinocytes [59], as well as in vivo recruitment of keratinocytes [72]. HMGB1 promotes angiogenesis by promoting the migration of endothelial cells [77]. HMGB1, HMGN1, EDN, uric acid, S100A8/9, cathelicidin, and HSPs have all been shown to be important for the induction of inflammation and immune responses [49, 81, 84, 101–106], which is likely contributed by the capacities of these alarmins to both mobilize and activate leukocytes. HMGN1-induced DC recruitment is responsible at least in part for the capacity of HMGN1 to induce immune responses, since HMGN1^−/− mice manifest remarkably reduced immune responses accompanied by greatly reduced recruitment of DCs to the site of immunization [49]. Thus the capacity of alarmins to induce the recruitment of immune cells is critical for their participation in host defense responses. Consequently, inhibition of the release or activity of HMGB1 can ameliorate many inflammatory conditions (reviewed recently [107]). For example, blockade of HMGB1 upon heart transplantation also reduces infiltration of leukocytes and anti-graft Th1 immune responses, and significantly prolongs allograft heart survival [108].

Perspective

Although much has been learned in the past decade about the capacities of alarmins to induce cell migration and recruitment, much more needs to be done in order to elucidate the intracellular signaling pathways utilized by various alarmins; of note, the receptors for many alarmins remain unidentified. It is also critical to dissect to what extent each alarmin contributes to the trafficking of diverse types of cells in vivo under distinct conditions. By further understanding the role alarmins play and the mechanisms involved, ways of targeting alarmin-induced trafficking of immune cells may be identified and this provide a means for regulating immune responses and treating inflammatory and autoimmune disorders.

Abbreviations:

CRAMP

cathelicidin-related antimicrobial peptide

DAMP

damage-associated molecular pattern

EDN

eosinophil-derived neurotoxin

EPC

endothelial progenitor cell

GiPCR

Gαi protein–coupled receptor

HMGB1

high-mobility group box-1 protein

HMGN1

high-mobility group nucleosome-binding protein 1

MC

mast cell

PI3K

phosphatidylinositol-3 kinase

RAGE

receptor for glycation end products

SMC

smooth muscle cell