Integrin binding

Sticking around vessels

A study demonstrates that controlled integrin binding on a biomaterial was capable of promoting vascular cell sprouting and formation of a non-leaky blood vessel network in a healthy and diseased state.

Blood vessel formation — neovascularization — is a critical process supporting tissue development, regeneration and tumorigenesis. It is therefore essential to understand how neovascularization is regulated in a tissue-specific context towards developing therapeutics. Elucidation of neovascularization reveals a complex process inherently dependent on the presence of growth factors and cytokines (bioactive factors), concurrent with dynamic regulation of the local extracellular matrix (ECM) properties, including matrix stiffness, degradability and binding motif presentation (Fig. 1a). Until now, inducing neovascularization with a range of approaches has met numerous challenges, including off-target effects, difficulties in dosing and delivery, and pathophysiological vascular regeneration defined by leaky, unorganized vessels. Therefore, the basic understanding of the process of neovascularization in a range of applications has generated a lot of interest. Recent research has demonstrated how surface receptors on cells are not only essential for the interaction of cells with their surrounding matrix but also for the formation of blood vessels. In this context, Tatiana Segura and colleagues1 now report in Nature Materials how blood vessel networks can be regulated by integrin activators presented on a biomaterial.

Figure 1: Integrin-binding effects on cell patterning and vascularization.
figure1

a, Without optimal bioactive cues (for example, growth factor) and intrinsic mechanical properties (for example, matrix rigidity), endothelial cells remain quiescent. In the presence of sufficient bioactive cues and ECM stiffness amenable to angiogenesis, tip cells extend into the matrix by metalloproteinase-mediated matrix degradation and cell–ECM interactions. Tip cells are followed by stalk cells to form neovessels. b,c, Segura and colleagues employed angiogenic sprouting beads to study vascular regeneration. Both integrin-stimulating materials (α3/α5β1 and αvβ3) allow for sprouting from beads. Sprouting into α3/α5β1-stimulating matrices leads to formation of organized, space-filling vasculature (b). Sprouting into αvβ3-stimulating matrices leads to disorganized, looping vasculature that fills a lower percentage of the surrounding matrix with new vessels (c). Anastomosis between sprouting beads is observed with both integrin stimulators, but α3/α5β1 matrices establish more organized junctions in vitro and permit formation of organized, non-leaky vessels in vivo; αvβ3 matrices form disorganized junctions between new vessels and leaky vasculature in vivo.

The significance of cell–ECM interaction has received a lot of attention, and this fascinating interplay has been investigated with a range of substrates. It has been shown that endothelial cell interaction with interstitial matrix proteins such as collagen is modulated by integrins, and this association is fundamental in the activation of endothelial cells and tubular morphogenesis2. Polymers lacking intrinsic cell-binding capacity provide a blank-slate approach and lend themselves to isolation and elucidation of the role of a number of biomaterial properties. A 'plug and play' approach can be applied with such biomaterials, and precise mechanistic questions can be answered. Peptides such as Arg-Gly-Asp (RGD), which can mimic integrin-binding sequences on naturally occurring proteins, can be incorporated within polymeric biomaterials to study, for instance, the sequential activation of vascular morphogenesis in an engineered matrix3. Using this platform, it has been shown that both integrins α5β1 and αvβ3 regulate vascular morphogenesis in a polymeric matrix. This tunable platform has been used to demonstrate the therapeutic potential of human-engineered microvasculature networks, using pathological in vivo model systems. Interestingly, the role of integrins appears much more complex than at first glance. Rather than acting just as cellular Velcro enabling cell–ECM adhesion, integrins represent an incredibly elegant collective means of cell–ECM communication, and also act as potent bioactive regulators that can activate transduction of signals that inform cell behaviour4.

Expanding this idea, Segura and colleagues utilize a biomaterial as a platform to present integrin activators to modulate endothelial cell patterning and vessel network formation in vitro and in vivo. Briefly, the extracellular glycoprotein, fibronectin, and its fragments were engineered to preferentially bind to two specific integrins, α3/α5β1 and αvβ3. Using the plug and play approach, the attachment and spreading of endothelial cells on surfaces was shown to be dependent on the type of integrins activated by the fibronectin fragments (Fig. 1b,c). In addition, the researchers also cultured endothelial cells on 'sprouting beads' in fibrin hydrogels and showed that the presence of native fibronectin in the hydrogel was crucial in endothelial cell sprouting and branching networks. The morphology of these sprouting networks was also found to be dependent on the type of integrins activated by protein fragments. The use of fibrin hydrogels containing depleted fibronectin reduced endothelial cell sprouting, although this effect could be rescued by the addition of exogenous fibronectin or RGD peptides.

An important process in neovascularization is the fusion of sprouting and pre-existing vessels to form a complete network of functional blood vessels in what is known as anastomosis. Activation of either α3/α5β1 or αvβ3 in fibrin hydrogels facilitated connections between vascular sprouts reminiscent of anastomosis. However, markers of anastomosis were found to be significantly altered on endothelial cells depending on the type of integrins activated, with αvβ3 stimulation leading to less organized connections. To confirm in vitro observations, two in vivo assessments were carried out using hyaluronic acid hydrogels encapsulated with fibronectin fragments to activate the two integrin types — α3/α5β1 or αvβ3 — as well as nanoparticles containing vascular endothelial growth factor. These hydrogels were first implanted into mice subcutaneously. The hydrogels with the α3/α5β1 integrin-specific fragment-stimulated formation of vasculature with organization and morphological characteristics similar to native blood vessels, whereas αvβ3 stimulating hydrogels led to disorganized, tortuous vessels. Another in vivo study was carried out using a mouse stroke model. Following the induced stroke, the hydrogels were injected into the stroke cavity in the brain and, as expected, the area covered by the neovascular network was greater in the presence of the α3/α5β1 integrin-specific fragment versus the αvβ3 fragment. Importantly, it was evident that α3/α5β1-induced vessels had a decreased leakage (Fig. 1b) or permeability. This leakiness is symptomatic of vessels grown in the presence of hyperphysiological concentrations of angiogenic growth factor proteins or vessels forming in pathophysiological conditions.

Ultimately, the findings of the current work further highlight the importance of understanding the specific and dynamic role of integrins throughout vascular regeneration and vessel stabilization in healthy and diseased states. Moving forward, a more in-depth understanding of the observed phenomena should be sought in terms of elucidating downstream effects of integrin stimulation related to vascular maturity. Researchers should also take into account the dynamic nature of the surrounding environment during the process of vessel formation, especially as the local ECM is remodelled through degradation and matrix deposition. Moreover, the balance of cell–cell and cell–ECM interactions can alter perception of matrix stiffness and influence cell proliferation and fate decisions5. All these factors may alter the role of integrins in neovascularization and should be the subject of further studies investigating the process of cell and tissue patterning.

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Correspondence to Sharon Gerecht.

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Blatchley, M., Gerecht, S. Sticking around vessels. Nature Mater 16, 881–883 (2017). https://doi.org/10.1038/nmat4977

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