Identification

The phenotypic nature of pericytes was largely disclosed by transmission and scanning electron microscopy. However, due to the morphological similarities of rodent retinal capillary endothelial cells and pericytes, markers allowing for the distinction of retinal capillary cells were crucial for quantitative analysis.

There is still no “pan pericyte marker” due to the versatility of pericytes, even in one organ such as the retina (27). Common markers of pericytes are smooth muscle actin (SMA), desmin, the proteoglycan NG2, and the platelet-derived growth factor-receptor beta (PDGFR-beta). The aminopeptidase N, the identification of the expression of XlacZ gene in pericytes (and vSMC), and the regulator of G-signaling 5 (RGS5) are also used for identification (14, 35–44). By comparing cDNA microarrays of mouse brain of PDGF-B knockout with wildtype embryos, several downregulated genes were expressed in brain capillary pericytes of wildtype tissue, such as the ATP-sensitive potassium channel complex (Kir 6.1), the sulfonylurea receptor 2 (SUR2) and the delta homolog 1 (DLK1) (45).

The XlacZ4 mouse is widely used as a model to study the role of pericytes and SMC because it expresses a reporter gene under control of a pericyte/SMC-specific promoter. However, according to our analysis, approximately 55–65% of retinal pericytes express the XlacZ4 and the expression is context dependent. Common to all markers mentioned is that they fail to recognize all pericytes at all stages (26).

FUNCTION

Contractility

Pericytes are the capillary counterparts to SMC on arterioles and arteries (44, 46). One remarkable feature that pericytes have in common with SMC is their contractile phenotype. Pericytes contain both smooth muscle and nonsmooth muscle isoforms of actin and myosin, however, with an uneven distribution within the pericyte population (39). The differential expression of SMA in pericytes may reflect the continuum from SMC of arteries and arterioles to pericytes of true capillaries and may correlate with the physical forces that pericytes are exposed to. The same pericytes are immunolabeled with smooth muscle tropomyosin and cGK suggestive of a contractile function (37, 47, 48). Meanwhile, several factors are identified that regulate pericytes contractility. While alpha 2-adrenergic agonists, cholinergic agonists, histamine, serotonin, angiotensin II, and endothelin-1 lead to vasoconstriction, beta-2 adrenergic agonists, NO, and atrial natriuretic peptide lead to a dilatation of the pericyte-covered capillaries (34). As demonstrated in in vivo studies, pericytes in brain and retinal capillaries constrict in response to electrical stimulation, superperfusion with ATP and noradrenalin. These data provide firm evidence that pericyte control capillary blood flow in response to local modulation by vasoactive mechanisms (49). However, whether the hyperglycemic milieu or the loss of pericytes has an impact on contractility, in particular when basement membrane components have changed under hyperglycaemic conditions, awaits further clarification.

Role in Vessel Formation and Stabilization

The function of pericytes that has been most intensively studied is their role in endothelial proliferation and angiogenesis. Studies also highlighted the importance of pericytes for vessel maturation in embryonic development, in vascular remodeling, and in guidance of sprouting angiogenesis (50–54). Endothelial cells alone can initiate, but not complete vessel formation. After the primary network of vessels has formed during vasculogenesis, maturation of the primitive network ensues. During this process, pericytes are recruited to the forming vasculature. Important molecular pathways, involved in pericyte recruitment during embryonic vessel maturation, are platelet-derived growth factor-beta (PDGF-B) and its receptor (PDGFR-B), transforming growth factor-beta (TGF-β) and its receptor and the angiopoietin/Tie-2 system (Ang/Tie-2). While the PDGF/PDGF-receptor system is crucial for pericyte migration and proliferation during vascular maturation, the angiopoietin/Tie-2 system is essential for subsequent vessel stabilization, and TGF-β is involved in the interaction of vascular cells with extracellular matrix (ECM) and ECM production and further mural cell differentiation. Inhibition of pericyte recruitment to capillaries by interfering with the recruitment leads to abnormal remodeling of developing vessels, a process that is reversed by administration of endothelial survival factors (55). During this angiogenic remodeling the initial vascular network is also modified through both pruning and vessel enlargement (56, 57).

In sprouting angiogenesis, pericytes are actively involved (56). Under the influence of VEGF, endothelial cells start to evade from their resident site toward a VEGF gradient (58). Endothelial cell proliferation and migration of sprouting tip cells include the degradation and losing of ECM and is dependent on tip cell guidance (59). In this process, pericytes are recruited to vessels by the platelet-derived growth factor PDGF-B/ PDGFR-B system (60), but other factors such as sphingosine-1-phosphate-1 (S1P-1) and the angiopoietins are also involved (61). For the completion of vessel maturation, active TGF-ß signaling via the ALK-1 and Smad5 pathway is needed (62).

Vessels are resistant to hyperoxic vasoregression when covered with pericytes. This led to the concept of the “window of plasticity” determined by pericyte recruitment lagging behind endothelial sprouting (63). By the use of complementary phase-specific pericyte markers outlined earlier, it was shown that pericytes play an active role in physiological and pathological angiogenesis, as they are frequently found near, at or even in front of the tips of endothelial sprouts (14, 27, 64). When remodeling ceases, pericytes contribute to the stabilization of vessels by the production of collagen and ECM proteins, such as fibronectin, laminin, and glycosaminglycans to the basal lamina (33, 65–68). Once the entire vascular system has formed, the major function of pericytes becomes the maintenance of a functional vascular network by their ability to control endothelial cell proliferation and vascular tone and by making endothelial cells refractory to shifts in oxygen tension and growth factor levels (69–72). Another paracrine signaling pathway implicated in vessel stabilization is the angiopoietin/Tie-2 system. Angiopoietins (Ang-1 and Ang-2) signal via the tyrosine kinase receptor Tie-2. Pericytes are a predominant source of Ang-1 (54, 73). Ang-1 in endothelial cells is believed to maintain vessel integrity by its ability to stabilize nascent vessels and to promote tightness of vessel barrier function, presumably by facilitating communication between ECs and pericytes (31).