Inflammation is a complex biological response that serves to protect the body's tissues following harmful stimuli such as infection, irritation or injury and initiates tissue repair. At the start of an inflammatory response, pro-inflammatory mediators induce changes in the endothelial lining of the blood vessels and in leukocytes. This results in increased vascular permeability and increased expression of adhesion proteins, and promotes adhesion of leukocytes, especially neutrophils to the endothelium. Adhesion is a prerequisite for neutrophil extravasation and chemoattractant-stimulated recruitment to inflammatory sites, where neutrophils phagocytose and kill microbes, release inflammatory mediators and cross-talk with other immune cells to co-ordinate the immune response in preparation for tissue repair. Many signalling proteins are critically involved in the complex signalling processes that underpin the inflammatory response and cross-talk between endothelium and leukocytes. As key regulators of cell–cell and cell–substratum adhesion, small GTPases (guanosine triphosphatases) act as important controls of neutrophil-endothelial cell interactions as well as neutrophil recruitment to sites of inflammation. Here, we summarise key processes that are dependent upon small GTPases in leukocytes during these early inflammatory events. We place a particular focus on the regulation of integrin-dependent events and their control by Rho and Rap family GTPases as well as their regulators during neutrophil adhesion, chemotaxis and recruitment.

Inflammation as an innate, immuno-vascular response

Classical inflammation is a complex immuno-vascular response that is triggered by harmful stimuli such as pathogens, tissue injury or irritants. Inflammation precedes and sets the foundation for healing. The induction of inflammation is triggered by the release of vasoactive pro-inflammatory mediators. These mediators trigger rapid vascular changes, including vessel dilation, decreased blood flow and increased vascular permeability [1,2]. The extent of these events is dependent upon the vascular bed, with e.g. high permeability in post-capillary venules and very little in brain vessels. These vascular changes permit blood plasma and antimicrobial proteins, such as complement factors and antibodies contained within it to enter the surrounding tissue, leading to oedema in a process that is referred to as ‘vascular leakage’ (Figure 1).

Vascular leakage and leukocyte–EC interactions in inflammation.

Figure 1.
Vascular leakage and leukocyte–EC interactions in inflammation.

With the induction of an inflammatory response, the endothelium is stimulated and remodelled by the cytokines released by tissue-resident immune cells such as macrophages, resulting in the up-regulation of P- and E-selectins and vascular leakage. At the same time, the leukocyte recruitment cascade is initiated which involves several sequential stages, tethering, rolling, firm adhesion and TEM. Circulating neutrophils are captured by the endothelium through the binding of P-selectin glycoprotein ligand 1 (PSGL-1) by the P- and E-selectins on the ECs. Loosely tethered neutrophils then start rolling on the endothelium, and the engagement of the selectins and glycoproteins induces inside-out signalling in the neutrophil, shifting the conformation of β2 integrins, initially αLβ2 /LFA1 and later αMβ2/Mac1 from inactive to intermediate with higher ligand-binding affinity to ICAM ligands expressed by the endothelium. This mediates crawling along, and subsequently firm adhesion of neutrophils to the endothelium. To reach the site of inflammation, neutrophils transmigrate across the endothelium in a process that requires leukocyte integrins and endothelial junctional proteins such as PECAM-1. The paracellular mechanism, where leukocytes use EC junctions is shown here. Please note that the selectin and integrin usage and dependency for leukocyte extravasation does not appear to be universal across all endothelial beds.

Figure 1.
Vascular leakage and leukocyte–EC interactions in inflammation.

With the induction of an inflammatory response, the endothelium is stimulated and remodelled by the cytokines released by tissue-resident immune cells such as macrophages, resulting in the up-regulation of P- and E-selectins and vascular leakage. At the same time, the leukocyte recruitment cascade is initiated which involves several sequential stages, tethering, rolling, firm adhesion and TEM. Circulating neutrophils are captured by the endothelium through the binding of P-selectin glycoprotein ligand 1 (PSGL-1) by the P- and E-selectins on the ECs. Loosely tethered neutrophils then start rolling on the endothelium, and the engagement of the selectins and glycoproteins induces inside-out signalling in the neutrophil, shifting the conformation of β2 integrins, initially αLβ2 /LFA1 and later αMβ2/Mac1 from inactive to intermediate with higher ligand-binding affinity to ICAM ligands expressed by the endothelium. This mediates crawling along, and subsequently firm adhesion of neutrophils to the endothelium. To reach the site of inflammation, neutrophils transmigrate across the endothelium in a process that requires leukocyte integrins and endothelial junctional proteins such as PECAM-1. The paracellular mechanism, where leukocytes use EC junctions is shown here. Please note that the selectin and integrin usage and dependency for leukocyte extravasation does not appear to be universal across all endothelial beds.

Pro-inflammatory mediators also activate circulating leukocytes. In the interest of space, we will focus this discussion on the neutrophil, a particularly important innate immune cell in early inflammation [3,4]. Slowed blood flow leads to leukocyte margination, initiating close mechanical contact with endothelial cells (ECs). Meanwhile, endothelial exposure to pro-inflammatory cytokines induces increased display of adhesion molecules and their ligands on the luminal side of the vessel and on the neutrophil (Figure 1), enabling the leukocyte adhesion cascade (reviewed in [5]). In a selectin and integrin-mediated process, leukocytes develop increasingly strong and long-lived interactions with ECs. Individual stages include selectin-mediated tethering and rolling, selectin and integrin-mediated slow rolling and crawling, until integrin-mediated firm adhesion of the leukocyte. The process culminates in integrin-dependent diapedesis (extravasation; Figure 1). Neutrophils breach several barriers, the ECs, basement membrane and pericytes. Of these, neutrophil transendothelial migration (TEM) is most studied. TEM can occur by two routes, through endothelial junctions (paracellular) or through EC bodies (transcellular), with paracellular TEM observed most frequently in areas with weak endothelial junctions [6]. Intravital imaging has indicated that leukocytes scan the endothelium and the underlying pericytes for transmigration sites, which are used repeatedly [7].

Many of the seminal findings in leukocyte–EC interactions were made by studying leukocyte adhesion in flow chambers in vitro, as well as by intravital imaging of post-capillary venules in the cremaster muscle, a site that is amenable to exteriorization and intravital imaging. With the advent of major advances in intravital imaging, other, less accessible vascular beds are also being analysed, e.g. lung and liver. A growing body of work indicates that specialisation of individual vascular beds dictates their requirement for individual adhesion proteins on both leukocyte and EC and for the occurrence of individual steps of the cascade (reviewed in [8]). For example, neutrophil recruitment in ICAM-1/P-selectin doubly deficient mice is affected in the peritoneum, but not in the lungs [9]. The current thinking is, therefore, that leukocyte–EC interactions are not uniform across all sites in the body.

Small guanosine triphosphatases

The Ras superfamily of small GTPases (guanosine triphosphatases) comprises several families, including Ras, Rho, Arf and Rab. Put very simply, Rap GTPases, which are part of the Ras family, regulate cell–cell and cell–substratum interactions, Rho GTPases are most famous for regulating dynamic actin rearrangements, while Arf and Rab small GTPases regulate intracellular transport. They are all required for complex cellular processes, such as single/collective cell migration, that are dependent on the actin cytoskeleton and the dynamic generation and dissolution of adhesive contacts [1012]. As such, small GTPases are key regulators of the neutrophil, a highly specialised cell that is able to change rapidly between circulating in the blood stream, adhering to the vessel wall under conditions of blood flow and migrating through tissue.

Small GTPases cycle between an active, GTP-bound and an inactive GDP-bound state (Figure 2). Small GTPases rely on their regulators, GTPase activating proteins (GAPs), guanine nucleotide exchange factors (GEFs) and GDP dissociation inhibitors (GDIs) [13]. GAPs increase the endogenous GTPase activity of the GTPase, inactivating it. In contrast, GEFs catalyse the exchange of GDP for GTP (guanosine triphosphate), thereby moving the small GTPase into the active state. Finally, GDIs sequester a subset of mostly Rho and Rab family small GTPases in the cytosol by shielding their lipid modification and protecting them from being activated. The number of Rho and Rap GEFs and GAPs outweighs that of the small GTPases themselves. These regulators fine-tune the precise timing and location of the GTPase activity. Each GEF/GAP is itself activated only under specific circumstances using defined upstream regulators at the right time and place [13]. Modes of regulation of small GTPases and their regulators are diverse, and can depend, among other mechanisms, on the formation of protein complexes, phosphorylation events and second messengers, such as the phosphoinositide 3-kinase (PI3K) lipid product phosphatidylinositol-(3,4,5)-trisphosphate [14].

The small GTPase cycle.

Figure 2.
The small GTPase cycle.

Small GTPases are molecular switches that regulate many cellular functions. They cycle between an active, GTP-bound (drawn in green) and an inactive, GDP-bound form (drawn in red). Guanosine phosphate groups are indicated by blue dots, and lipid modifications are indicated in brown. Guanine nucleotide exchange on small GTPases is catalysed by GEFs. The frequently slow intrinsic GTPase activity of small GTPases is activated by GAPs. Some small GTPases (especially Rho and Rab families) are in addition regulated by GDIs. GDI-dependent sequestration of the lipid modification of the small GTPase in the cytoplasm prevents degradation, nucleotide exchange and membrane association. Both GEFs and GAPs are subject to regulation by a variety of mechanisms, including, phosphorylation, second messengers and formation of protein complexes. Together, these mechanisms achieve the correct spatiotemporal activation of small GTPases in any given situation.

Figure 2.
The small GTPase cycle.

Small GTPases are molecular switches that regulate many cellular functions. They cycle between an active, GTP-bound (drawn in green) and an inactive, GDP-bound form (drawn in red). Guanosine phosphate groups are indicated by blue dots, and lipid modifications are indicated in brown. Guanine nucleotide exchange on small GTPases is catalysed by GEFs. The frequently slow intrinsic GTPase activity of small GTPases is activated by GAPs. Some small GTPases (especially Rho and Rab families) are in addition regulated by GDIs. GDI-dependent sequestration of the lipid modification of the small GTPase in the cytoplasm prevents degradation, nucleotide exchange and membrane association. Both GEFs and GAPs are subject to regulation by a variety of mechanisms, including, phosphorylation, second messengers and formation of protein complexes. Together, these mechanisms achieve the correct spatiotemporal activation of small GTPases in any given situation.

Small GTPases and neutrophils

Neutrophil biology and their regulation in health and disease have been the subject of several excellent recent reviews (e.g. [3,4]). Neutrophils are terminally differentiated, short-lived abundant circulating innate immune cells. These highly specialised leukocytes chemotax (move towards a chemoattractant) to sites of inflammation with exquisite speed and directionality. They efficiently and quickly phagocytose opsonised and non-opsonised bacteria and yeasts, and produce reactive oxygen species and degranulate (releasing a range of potent proteases and cytotoxic compounds that are stored in their specialised granules) to kill the ingested microbes inside the phagosome. These dynamic functions are all subject to regulation by small GTPases (Table 1 and [1518]). In the following, we concentrate on the function of Rho and Rap small GTPases in leukocyte–EC interactions.

Table 1
Neutrophils are subject to regulation by multiple small GTPases

Neutrophils carry out a range of specialised functions which allow them to ingest and kill pathogens and to generate inflammation. These functions are regulated by small GTPases as well as their regulators (not shown here). Please note that regulators identified in other model systems (e.g. macrophages in the case of phagocytosis) have not been included in this table. We apologise to the authors of many primary research papers that could not be cited here.

Neutrophil function Small GTPase involved References 
Adhesion Rac2 (under flow) [53
RhoA [68
Rap [69
Arf6 [23
Spreading Rac2 [53
Rap1 [32
Polarisation Cdc42 [70,71
Rac2 [72
RhoG [73
Rap1 [69
Chemotaxis Rac1, Rac2 [53,72
Cdc42 [70,71
RhoA [68
Rap1b [69,74
Rab27 [75
Arf6 [23,76
Recruitment Rac1, Rac2 [53,77
RhoA [68
Rap1b [74
Rab27 [75,78
Arf6 [76
Phagocytosis Arf6 [23
Rab5a [79
Rap1 [32
NADPH Oxidase Rac2 (but not Rac1) [53,77
RhoG [80
Rap1 [32
Arf6 [81,82
Rab27 [83
Degranulation Arf6 [81
Rab27 [83
Apoptosis RhoG [80
Cdc42 [84
NET release Rac2 [85
Rab27 [86
Neutrophil function Small GTPase involved References 
Adhesion Rac2 (under flow) [53
RhoA [68
Rap [69
Arf6 [23
Spreading Rac2 [53
Rap1 [32
Polarisation Cdc42 [70,71
Rac2 [72
RhoG [73
Rap1 [69
Chemotaxis Rac1, Rac2 [53,72
Cdc42 [70,71
RhoA [68
Rap1b [69,74
Rab27 [75
Arf6 [23,76
Recruitment Rac1, Rac2 [53,77
RhoA [68
Rap1b [74
Rab27 [75,78
Arf6 [76
Phagocytosis Arf6 [23
Rab5a [79
Rap1 [32
NADPH Oxidase Rac2 (but not Rac1) [53,77
RhoG [80
Rap1 [32
Arf6 [81,82
Rab27 [83
Degranulation Arf6 [81
Rab27 [83
Apoptosis RhoG [80
Cdc42 [84
NET release Rac2 [85
Rab27 [86

Integrins

Integrins are extracellular receptors composed of two chains (α/β) that are expressed by all nucleated cells [19]. Using their extracellular domains, integrins bind to their ligands. Whereas integrin ligands are normally extracellular matrix proteins, ligands of leukocyte integrins are often expressed on the surface of other cells, e.g. endothelial ICAMs and VCAMs. Integrin intracellular tails form dynamic links to the cellular actin cytoskeleton. In this way, integrins mediate cell adhesion to anchor cells and to partake in dynamic cellular functions requiring such contacts, such as cell migration.

Integrins signal bidirectionally [19] with ligand-binding-induced signalling referred to as ‘outside-in signalling’, whereas ‘inside-out signalling’ refers to intracellular signalling that regulates integrin activation. This allows inactive integrins in their bent conformation to adopt intermediate and finally the active (extended) conformations (see Figure 3 for a simplified drawing). Mechanistically, interactions with intracellular activators, notably talin and kindlin, convert the inactive integrin to the active conformation [20]. Integrin ligand-binding activity is furthermore regulated by clustering, which promotes ligand-binding avidity. The regulation of the integrin ligand-binding avidity and affinity states is tightly interconnected, although the precise molecular mechanism underpinning integrin avidity remains controversial. Integrins are subject to dynamic internalisation and recycling events, in addition to the regulation of ligand-binding activity, that are regulated by many different pathways which employ Rab and/or Arf family small GTPases [12]. Our understanding of the regulation of integrin trafficking is currently growing at a rapid pace, with experiments typically performed in cultured adherent cell lines. Intracellular trafficking events are at times dependent on integrin receptor and also cell line. Neutrophils are known to express numerous Rab and Arf GTPases [21,22]. The Arf6 GEF cytohesin1 has been reported to regulate Mac-1 in neutrophils [23], but for the most part, specific roles of Arf/Rab-dependent integrin trafficking remain to be elucidated in the neutrophil.

Rap-dependent regulation of integrin inside-out signalling.

Figure 3.
Rap-dependent regulation of integrin inside-out signalling.

The shift from the inactive (bent) to the active (extended) conformation is regulated by integrin inside-out signalling. Talin (and kindlin, not shown) binding to the cytoplasmic tail of the integrin β-subunit is required for integrin activation, and regulated by the small GTPase Rap in combination with its effectors, the best understood of which is RIAM. The formation of a Rap-RIAM-talin complex that binds integrin mediates integrin activation. Several GEFs upstream of Rap have been identified in integrin inside-out signalling, of which CalDAG-GEF1 is thought to be particularly important in neutrophils. Two negative regulators have been described that interfere with Rap/talin-mediated integrin activation. SHANK proteins can sequester Rap and interfere with the formation of the Rap-RIAM-talin complex, whereas SHARPIN binding to the integrin α-subunit cytoplasmic tail interferes with the complex binding to the β-subunit.

Figure 3.
Rap-dependent regulation of integrin inside-out signalling.

The shift from the inactive (bent) to the active (extended) conformation is regulated by integrin inside-out signalling. Talin (and kindlin, not shown) binding to the cytoplasmic tail of the integrin β-subunit is required for integrin activation, and regulated by the small GTPase Rap in combination with its effectors, the best understood of which is RIAM. The formation of a Rap-RIAM-talin complex that binds integrin mediates integrin activation. Several GEFs upstream of Rap have been identified in integrin inside-out signalling, of which CalDAG-GEF1 is thought to be particularly important in neutrophils. Two negative regulators have been described that interfere with Rap/talin-mediated integrin activation. SHANK proteins can sequester Rap and interfere with the formation of the Rap-RIAM-talin complex, whereas SHARPIN binding to the integrin α-subunit cytoplasmic tail interferes with the complex binding to the β-subunit.

The β2 integrins αMβ2 (also known as CD11b/CD18, Mac1 or CR3) and αLβ2 (also known as CD11a/CD18 or LFA1) are the major neutrophil integrins, but neutrophils also express other leukocyte integrins such as α4β1 (CD49d/CD29; VLA4) as well as RGD-binding integrins, e.g. α5β1 (CD49e/CD29, fibronectin receptor) αvβ3 (CD51/CD61, vitronectin receptor). β2 integrin function is probably best characterised in neutrophil firm adhesion to endothelial ICAM-1 and TEM in post-capillary venules, thanks to a large body of work by many investigators who employed blocking and/or affinity status-specific antibodies as well as knock-out mice as well as neutrophils from leukocyte adhesion deficiency (LAD) patients. In recent years, the use of ever advancing imaging techniques has helped to visualise these processes in increasingly fine detail. Such investigations have built onto older observations to illustrate how neutrophils flatten their bodies [24,25] and employ catch-bonds [26], long tethers [24] and slings [27] to enable and stabilise rolling even under conditions of high sheer stress. In contrast, neutrophil recruitment to the alveoli of the inflamed lung, and extravascular migration have been shown to depend on β1 integrins (e.g. [28,29]). α5β1 and αLβ2 integrins were shown to be subject to recycling from the rear towards the front of the migrating neutrophil [30,31].

Small GTPases in the regulation of (leukocyte) integrins

Rap small GTPases represent the best-understood enzymatic regulators of the activity status of integrins. Rap1A-deficient mice are characterised by integrin-dependent leukocyte adhesion defects [32,33]. Three distinct Rap effectors, RAPL [34], RIAM [35] and RADIL [36] have all been shown to function as links to integrin affinity and avidity. The mechanism involving RIAM-mediated integrin regulation is best characterised. The formation of Rap1–RIAM–talin complexes allows the recruitment of talin to integrin, inducing integrin activation [37]. Genetic experiments indicate that RIAM is particularly important for the activation of leukocyte β2 integrins [38,39], while alternative mechanisms appear to operate in other cell types such as platelets [40]. Several Rap GEFs were shown to be involved in integrin activation. The cAMP-activated Rap GEFs Epac1/2 [4143] were shown to regulate integrin-mediated adhesion in many cell types, including in some leukocytes (e.g. [4446]). Although expressed by neutrophils, Epac activation is not thought to be sufficient for neutrophil integrin activation [47]. A rare mutation in CalDAG GEF1 (also known as RasGRP2) was found to result in LAD type III [48], and neutrophils from CalDAG GEF1-deficient mice have adhesion and recruitment defects in keeping with this disease [49].

The Rho GTPases, RhoA, Rac and Cdc42, are also involved in dynamic cell–matrix adhesion [50]. The function of Rho GTPases in integrin outside-in signalling has been clearly documented (reviewed in [51]), but there is little evidence for Rho-dependent integrin inside-out signalling from experimentation with cultured adherent cell lines. In contrast, analyses of leukocytes under flow conditions have shown that Rap and Rho small GTPases are involved in the activation of leukocyte integrins (Figure 4). The first leukocyte–EC interaction, neutrophil capture, relies on display of stored endothelial P-selectin. This is followed by rolling mediated by endothelial P-selectin/E-selectin and leukocyte L-selectin each binding their carbohydrate ligands on their counterparts. Leukocyte rolling induces a shift in the leukocyte integrin αLβ2 activation to an intermediate state, which, in turn, promotes slow rolling. Chemokines encountered during rolling induce further leukocyte integrin activation to the fully extended conformation. Cal-DAG-GEF1-mediated Rap1A activation was shown to be required for slow rolling of neutrophils to occur, due to Rap's role in rolling-mediated leukocyte integrin activation [52]. Rac2-deficient mouse neutrophils, and those isolated from a patient who carried a dominant negative Rac2 mutation were characterised by defective rolling on P-selectin [53,54]. It remains unclear whether this defect might have been secondary to a defect in conveying αLβ2 activation. Neutrophils deficient in the Rac GEF P-Rex1 also displayed a defect in slow rolling, which was shown to be due to a role of P-Rex1 in the selectin-mediated activation of leukocyte integrins [55]. Intravital imaging of airway post-capillary venules in inflamed lungs suggested that the Rac GEFs P-Rex1 and Vav GEFs together regulate Rac-mediated αLβ2-activation in this context [56]. RhoA and Rac were also shown to mediate rapid chemokine-induced αLβ2 activation (inside-out signalling) in T lymphocytes under flow conditions [57,58]. In contrast with neutrophils, primary T cells are amenable to being cultured and transfected. Careful analysis of signalling events in primary human lymphocytes identified CXCL12-induced αLβ2 inside-out signalling involved GαI-JAK2/3-Vav1-Rho-PLD1-Rap1 signalling [59,60]. Additional knock-down studies with primary human lymphocytes identified that further Rho GEFs, SOS1, ArhGEF1 and DOCK2 are also involved in the chemokine-mediated affinity regulation of αLβ2 [61]. In contrast, Cdc42 was shown to counteract chemokine-driven leukocyte integrin activation under flow [59] and to interfere with chemoattractant-driven Rap activation [62].

Regulation of leukocyte integrin inside-out signalling by Rho GTPases.

Figure 4.
Regulation of leukocyte integrin inside-out signalling by Rho GTPases.

Chemokine-driven integrin activation in the leukocyte adhesion cascade depends on Rho GTPase signalling in neutrophils and thymocytes. In thymocytes, CXCL12 binding causes Vav (and other) RhoGEF activation, inducing RhoA activation. This drives Rap activation (and Rap-mediated integrin activation) by making use of an indirect mechanism involving PLD1 and an as-yet-undefined Rap GEF.

Figure 4.
Regulation of leukocyte integrin inside-out signalling by Rho GTPases.

Chemokine-driven integrin activation in the leukocyte adhesion cascade depends on Rho GTPase signalling in neutrophils and thymocytes. In thymocytes, CXCL12 binding causes Vav (and other) RhoGEF activation, inducing RhoA activation. This drives Rap activation (and Rap-mediated integrin activation) by making use of an indirect mechanism involving PLD1 and an as-yet-undefined Rap GEF.

As is often the case with switching off biological processes, our understanding of integrin inactivation lags far behind that of integrin activation. It stands to reason that such mechanisms not only exist, but, given the importance of integrin signalling, will be subject to tight regulation. Neutrophils from mice deficient in the PI3K and Rap-regulated RhoA and Arf6 GAP ARAP3, or those in which ARAP3 was uncoupled from activation by PI3K, were characterised by increased β2 integrin ligand binding, by increased adhesion under static and flow conditions and by increased outside-in signalling in vitro. In vivo, these neutrophils were characterised by reduced crawling and increased firm adhesion and by a recruitment defect in sterile inflammation in the context of bone marrow chimeras [63,64]. Collectively, this is suggestive of a function of small GTPases also in the regulation of integrin inactivation. Moreover, two mechanisms that compete with integrin activation have recently been described (Figure 3). First, SHARPIN binding to integrin α subunits was shown to interfere with talin and kindlin binding to β-subunits and integrin activation [65]. SHARPIN was subsequently shown to bind the α subunit of lymphocyte αLβ2, and to colocalise with αLβ2 to the trailing end of migrating T lymphocytes; in vivo, Sharpin-deficient T lymphocytes were deficient in homing to lymph nodes [66]. Second, sequestration of GTP-Rap by SHANK1/3 proteins has recently been shown to interfere with Rap-RIAM-talin-mediated integrin activation in cancer cells, promoting the inactive integrin state ([67]; Figure 3).

Further information on small GTPase-dependent functions in the regulation of integrins in inflammation is bound to emerge in the future. It will be exciting to watch this space.

Abbreviations

     
  • EC

    endothelial cell

  •  
  • GAP

    GTPase activating protein

  •  
  • GDI

    GDP dissociation inhibitor

  •  
  • GEF

    guanine nucleotide exchange factor

  •  
  • GTP

    guanosine triphosphate

  •  
  • GTPase

    guanosine triphosphatase

  •  
  • LAD

    leukocyte adhesion deficiency

  •  
  • PI3K

    phosphoinositide 3-kinase

  •  
  • TEM

    transendothelial migration

Author Contribution

J.C. and S.V. wrote the initial version and drafted the figures; all authors wrote the final version of the paper.

Funding

This work was funded by the Medical Research Council [MR/K501293/1] and by the British Heart Foundation [PG/17/54/32981].

Competing Interests

The Authors declare that there are no competing interests associated with the manuscript.

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