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Endocr Rev. Jun 2010; 31(3): 343–363.
Published online Feb 17, 2010. doi:  10.1210/er.2009-0035
PMCID: PMC3365844

The Role of Blood Vessels, Endothelial Cells, and Vascular Pericytes in Insulin Secretion and Peripheral Insulin Action

Abstract

The pathogenesis of type 2 diabetes is intimately intertwined with the vasculature. Insulin must efficiently enter the bloodstream from pancreatic β-cells, circulate throughout the body, and efficiently exit the bloodstream to reach target tissues and mediate its effects. Defects in the vasculature of pancreatic islets can lead to diabetic phenotypes. Similarly, insulin resistance is accompanied by defects in the vasculature of skeletal muscle, which ultimately reduce the ability of insulin and nutrients to reach myocytes. An underappreciated participant in these processes is the vascular pericyte. Pericytes, the smooth muscle-like cells lining the outsides of blood vessels throughout the body, have not been directly implicated in insulin secretion or peripheral insulin delivery. Here, we review the role of the vasculature in insulin secretion, islet function, and peripheral insulin delivery, and highlight a potential role for the vascular pericyte in these processes.

Abstract

Insulin transport from the bloodstream to its target cells requires transport across a vascular endothelial barrier. This step is regulated by many factors, including pericytes. Similarly, insulin transport from β-cells to the bloodstream requires efficient access to the vasculature. We review the role of the vasculature in insulin action and insulin secretion.

  • I. Introduction
  • II. Endothelial Cells and the Heterogeneity of Vascular Beds
  • III. Islet Vasculature and Insulin Secretion
    • A. Introduction to islet vasculature
    • B. The importance of the vasculature for pancreas development
    • C. Proper vascularization is also required for mature islet function
    • D. The role of islet revascularization during islet transplantation
  • IV. Peripheral Vasculature and Insulin Delivery
    • A. Introduction to peripheral vasculature and insulin delivery
    • B. Transendothelial transport of insulin
    • C. Effects of insulin on blood flow
    • D. Insulin-induced capillary recruitment
    • E. Molecular mechanism of capillary recruitment
    • F. Insulin resistance and muscle vasculature
    • G. Exercise-induced vascular changes
  • V. Vascular Pericytes: More Than Inert Contractile Cells
    • A. Introduction to pericytes
    • B. Platelet-derived growth factor-B: a key mediator of pericyte function
    • C. Diabetic complications: a key role for pericytes
    • D. Are pericytes multipotent progenitor cells?
    • E. Pericytes in normal islet function
    • F. Pericytes in islet tumors
    • G. A role for PDGF-B signaling in glucose uptake?
    • H. Inhibition of PDGFRβ and diabetes therapy
    • I. A role for pericytes in insulin-induced hemodynamic changes
  • VI. Summary/Conclusions

I. Introduction

Type 2 diabetes is a growing world epidemic (1,2). There appear to be two key steps in the development of type 2 diabetes: 1) the development of insulin resistance; and 2) β-cell decompensation. Although both of these processes are beginning to be understood at the molecular level, much remains to be elucidated. An important recent development is the discovery of the role that blood vessels play in the pathogenesis of these two conditions. The focus of this review is an investigation of the role that blood vessels and their constituent endothelial cells, vascular smooth muscle cells (vSMCs), and pericytes play in β-cell function and the development of insulin resistance. Several excellent reviews have described several of these topics (3,4), but this review will have a broader focus including both the role of blood vessels in islet development and function and introducing the pericyte as a novel mediator of these effects.

II. Endothelial Cells and the Heterogeneity of Vascular Beds

Blood vessels in the vascular beds of different tissues exhibit large structural variability, especially in the number of fenestrae and caveolae (5,6,7,8). Fenestrae are the approximately 100-nm pores covered by a permeable diaphragm resulting from the fusion of apical and basolateral plasma membranes. Caveolae are the 60- to 80-nm plasma membrane pits thought to be involved in endocytosis and transcytosis. For example, the highly permeable liver endothelium is termed “discontinuous” and contains larger than normal fenestrae that lack diaphragms (5). Liver endothelium also has many intercellular gaps that allow for easy access of blood-borne molecules to hepatocytes (5). In contrast, the nonfenestrated, caveolae-free endothelium of the brain vasculature contains many tight junctions and has very low permeability (5). This helps to form the blood-brain barrier, which regulates the entry of blood-borne molecules into the brain and preserves ionic homeostasis (9). The permeability characteristics of pancreatic islet and muscle vasculature lie somewhere between these two extremes. Islet vasculature is relatively permeable, and although it does not have gaps between endothelial cells, the endothelial cells are highly fenestrated to allow for facile nutrient sampling from blood, enabling islets to respond quickly to fluctuations in blood glucose and adjust insulin secretion as needed (10,11). In contrast, both cardiac and skeletal muscle vasculature are relatively impermeable to blood-borne macromolecules (5,6,12).

Endothelial cells form the inner lining of blood vessels. They are critically involved in many physiological functions, including control of vasomotor tone, blood cell trafficking, hemostatic balance, permeability, proliferation, survival, and immunity (5). However, endothelial cells are not the only component of blood vessels. Pericytes and vSMCs line the outer walls of blood vessels and play critical roles in blood vessel and endothelial cell function. Interestingly, despite the recent interest in the role of blood vessels in insulin secretion and insulin action, a role for pericytes in these processes has not been well described.

III. Islet Vasculature and Insulin Secretion

A. Introduction to islet vasculature

A complete understanding of the role of the vasculature in islet function requires an understanding of islet vascular architecture. Pancreatic islets are highly vascularized by a dense network of capillaries. The islet capillary network is approximately five times denser in islets than in the surrounding exocrine tissue (11). Although pancreatic islets account for approximately 1% of total pancreatic mass, they receive approximately 7–10% of total pancreatic blood flow (13). Traditionally, blood has been thought to flow from one to three arterioles into the intraislet capillary network and empty into venules in the islet periphery (14). However, real-time fluorescent tracing of blood flow patterns in mouse islets demonstrates that in the majority of cases, islets exhibit an inner-to-outer flow pattern where capillaries perfuse β-cells before other islet cell types and larger vessels exhibit an efferent rather than afferent flow pattern (15). β-Cells are intimately associated with islet endothelial cells and are usually no more than one cell distant from the bloodstream (16). Islet capillaries are highly fenestrated (10,11); in fact, they contain about 10 times more fenestrae than the surrounding exocrine tissue (8,11). This presumably allows for delivery of nutrients and growth factors into islets and enables efficient sampling of the blood to allow for rapid glucose-sensing.

Islet blood flow can be regulated by a number of metabolic and nonmetabolic factors. For instance, glucose almost doubles islet blood flow (17). Insulin decreases islet blood flow, likely due to resultant hypoglycemia, rather than a direct effect of hyperinsulinemia (18). ATP signals through islet A1 adenosine receptors to increase islet blood flow (19). This is in agreement with the effects of glucose on islet blood flow and suggests that increased islet cell metabolism promotes increased islet blood flow. Other nutrients and hormones have indirect effects on islet blood flow by interacting with vasodilators and vasoconstrictors (20,21). Finally, the central nervous system impacts islet perfusion; brain stem-dead donors have impaired islet hormone secretion (22). These studies demonstrate that islet blood flow is carefully controlled by various nutrients and growth factors and is exquisitely intertwined with the metabolic state of the organism.

B. The importance of the vasculature for pancreas development

A close relationship between islet endocrine cells and endothelial cells has been described, from early islet development through mature islet function. In the developing embryo, the pancreatic endoderm is located immediately beside the dorsal aorta, and the presence of the dorsal aorta is required for pancreatic differentiation and insulin expression (23). Overexpression of vascular endothelial growth factor-A (Vegf-a) in the developing pancreas under control of the pancreatic and duodenal homeobox 1 (PDX1) promoter leads to a hypervascularized pancreas with a 3-fold increase in islet number and islet area as well as ectopic insulin expression in the developing gut, suggesting that the vasculature is not only critical for initiation of pancreas development but also remains important for islet growth throughout maturity (23). The transcription factor PTF1A was identified as a crucial signal that is induced by signals from aortic endothelial cells and promotes the budding of the PDX1-positive pancreatic endoderm as well as insulin and glucagon expression (24).

Subsequent to the early requirement for pancreatic budding, endothelial cells, both directly and indirectly through sphingosine-1-phosphate, promote growth and budding of the dorsal pancreatic endoderm by induction of mesenchymal cell proliferation (25,26). This close relationship between islet endothelial cells and endocrine cells continues during the period of postnatal islet expansion. During the pronounced expansion of islet endocrine cells 1 wk after birth, there is an even more pronounced expansion of islet endothelial cells, leading to a marked increase in intraislet vascular density (27). Overexpression of VEGF-A by a tetracycline-inducible pancreas-specific bitransgenic system demonstrates increased islet endothelial cell formation. However, this actually causes reduced postnatal β-cell mass, suggesting that islet angiogenic signals must be maintained within narrow limits (28). These data indicate that endothelial cues are critically important for initiation of pancreatic budding and that endothelial cells remain vitally important for pancreas and islet growth throughout later embryonic and early postnatal development.

C. Proper vascularization is also required for mature islet function

The close relationship between islet endocrine cells and endothelial cells is important not only for pancreas development, but also for mature islet function. Knockout of VEGF-A by Pdx1-promoter-driven Cre recombinase leads to severely reduced islet vascular density, resulting in glucose intolerance (29). Interestingly, these mice display loss of endothelial fenestrae and increased numbers of endothelial caveolae, suggesting that caveolae attempt to compensate for the loss of secretory capacity that is brought about by the “tightening” of the remaining blood vessels (29). It is somewhat surprising that these mice do not display more severe glucose homeostasis defects given the severe reduction of islet vasculature. Additionally, no loss of islet cell area was noted despite the evidence linking endothelial cell cues to islet proliferation, as described above in Section III. B.

Several studies directly examined the role of VEGF-A in adult β-cell function. Because Pdx1 is expressed at high levels throughout the developing pancreas and at lower levels in adult β-cells, these studies used rat insulin promoter (RIP)-driven CRE to conditionally knock out VEGF-A specifically in β-cells. Similar to the PDX1-CRE knockout mice, the RIP-Cre VEGF-A knockouts displayed impaired glucose tolerance and defective in vivo insulin secretion (30,31). A role for VEGF-A in the adult islet has also been directly tested: tamoxifen-inducible PDX-1-CRE VEGF-A knockouts show glucose intolerance and reduced vessel density, although the effects of VEGF-A loss are less severe than when VEGF-A is absent from the beginning of pancreatic development (32). These studies suggest that loss of islet endothelial cells can negatively affect insulin secretion and contribute to glucose intolerance.

Not surprisingly, loss of islet endothelial cells has a more severe effect in obese animals. RIP-CRE-mediated deletion of VEGF-A in high-fat diet-fed animals leads to more severe glucose intolerance and reduction in insulin secretion than in lean knockouts (33). Interestingly, these high-fat diet-fed mice are able to normally expand their β-cell mass, suggesting that normal islet endothelial cell density is dispensable for compensatory islet hyperplasia in response to obesity. Two potential explanations for the preserved ability of these mice to expand their β-cell mass could be residual VEGF-A due to inefficiency of the Cre promoter, which can vary with the distance between loxP sites (34), or the presence of other VEGF isoforms in islets (35). Because Cre is usually not expressed in 100% of its target cells, the remaining cells could produce enough VEGF-A to explain the continued presence of some islet endothelial cells. These remaining endothelial cells could be sufficient to promote β-cell mass expansion.

In addition to VEGF-A, several other islet proteins have been demonstrated to play a role in adult islet vascularization and function. Fyn-related kinase, when overexpressed under control of the RIP, causes reduced in vivo insulin secretion and mild glucose intolerance, which appears to be due to reduced islet blood flow and abnormal capillary morphology (36). Similarly, β-cell or pancreas-specific deletion of von Hippel-Lindau factor, the protein that controls the degradation of hypoxia-inducible factor-1α (HIF-1α), leads to glucose intolerance and impaired insulin secretion, which is mediated through an increase in Hif-1α expression (37). This study demonstrates that a direct effect of vessel loss (i.e., increased hypoxia) is able to mediate the same effect as actual vessel loss. This might seem somewhat paradoxical, because HIF-1α directly up-regulates Vegf-a expression and thus would be expected to increase islet vascularization (38). However, HIF-1α also directly alters the expression of genes involved in β-cell function in these mice, reducing the capacity of β-cells to mediate glucose uptake, glucose metabolism, and insulin secretion (37). Conversely, mice in which the antiangiogenic factor thrombospondin-1 is knocked out have enlarged hypervascular islets (39). Taken together, these data suggest that decreased islet vascularization or blood flow can have a deleterious effect on islet insulin secretion and whole-body glucose tolerance. Conversely, increased islet vascularization has the opposite effect.

In addition to the data gleaned from various knockout mice, several animal models of diabetes suggest a close relationship between islet vascularization and insulin secretion. For example, in Zucker fatty (ZF) and Zucker diabetic fatty rats, islets become more vascularized during the obesity-induced expansion of islet mass (40). Interestingly, as diabetes and loss of β-cell mass ensue in the Zucker diabetic fatty rats, islet vasculature decreases (40). A similar association between loss of islet capillary density and progression of diabetes is observed in the Otsuka-Long-Evans-Tokushima fatty rat model (41). These data provide further evidence that islet mass and the amount of islet vasculature are critically linked. Similarly, islet blood flow and blood pressure are elevated in nonobese diabetic GK rats, obese Zucker rats, obese Wistar rats, and GK-Wistar F1 hybrid rats (42,43,44). Islet blood flow is similarly increased in 1-month-old ob/ob vs. lean B6 mice during a period of hyperglycemia, hyperinsulinemia, and β-cell expansion (45). Islet blood flow was normalized in 6- to 7-month-old ob/ob mice, suggesting that increased islet blood flow is important during expansion of β-cell mass in response to hyperglycemia. A number of islet capillary changes were noted in adult db/db mice, including loss of islet capillaries, increased capillary diameter, and pericyte hypertrophy (46). Together, these data suggest that an increase in islet vascularization and blood flow accompanies compensatory β-cell mass expansion in response to hyperglycemia. When β-cells are no longer able to expand and decompensation occurs, loss of islet vasculature also occurs. Interestingly, insulin-deficient mice have more islet capillaries and bigger islets (47), which could indicate that insulin inhibits islet vascularization. Alternatively, this effect may be a compensatory response whereby islet vasculature and islet size increase in response to insufficient levels of insulin secretion.

Islet endothelial cells have recently been shown to play a critical role in producing islet basement membrane. β-Cells appear incapable of forming their own basement membrane (48), and deletion of VEGF-A under the Pdx1 promoter results in a loss of islet, but not acinar tissue basement membrane (49). Vascular endothelium-produced laminin-411 and laminin-511 appear to be critical for insulin gene expression and β-cell proliferation in a β1-integrin-dependent manner (49). Similarly, treatment of islets with endothelium-conditioned culture medium increases glucose-stimulated insulin secretion and islet insulin content, an effect that is blocked by addition of a neutralizing antibody to the β1-chain of laminin (50). Purified islet endothelial cells can also stimulate β-cell proliferation through secretion of hepatocyte growth factor (51). These results might shed some light on the described link between islet hyperplasia and increased islet vascularization. Hyperglycemia could trigger VEGF-A secretion from islet endocrine cells. This in turn stimulates vessel growth, basement membrane production, and hepatocyte growth factor production and secretion, which ultimately leads to increased insulin production and β-cell proliferation. Some caution is necessary in applying these findings to human islets, however, because it was recently demonstrated that blood vessels in human islets are surrounded by a unique double basement membrane (52).

D. The role of islet revascularization during islet transplantation

One area in which the process of islet vascularization is thought to be especially important is during islet transplantation. The success of islet transplantation can critically hinge on the ability of transplanted islets to establish functional vasculature (53,54). In addition to immunorejection of newly transplanted islets, islet survival has proved a major challenge to the success of this procedure (55). Isolation of islets for transplantation damages islet endothelial cells and obviously involves the severing of native vasculature (56,57,58). Thus, newly transplanted islets must reestablish a functional vascular network, a process that is believed to involve angiogenesis and possibly vasculogenesis (57). Not surprisingly, newly transplanted islets are less vascularized immediately after transplant, and even after several weeks they remain less vascularized and maintain a lower oxygen tension than native pancreatic islets (59,60). Islet death and apoptosis are common problems after transplant, with islet cell apoptosis increasing and β-cell mass decreasing 1–3 d after transplant (61,62). Given the important connection between islet endocrine cells and islet endothelial cells described in Section III. C, it is not hard to imagine that lack of a functional vasculature might play a direct role in this increased islet death. Additionally, the resultant hypoxia and/or ischemia could also strongly contribute to increased islet apoptosis. In support of this, Hif-1α expression is increased in newly transplanted islets, and suppression of Hif-1α in transplanted islets reduces β-cell death (63). Once again, this might seem paradoxical, due to the ability of HIF-1α to increase Vegf-a expression (38), but HIF-1α also stimulates many other responses, including hypoxia-induced growth arrest and apoptosis, both of which would negatively impact the survival of transplanted islets (64).

Endothelial cells from the transplant recipient were originally thought to be the major contributor to posttransplantation vessel formation, but recent evidence suggests that donor endothelial cells also play an important role in islet revascularization. By using lacZ- or GFP-tagged donor endothelial cells, it has been demonstrated that after transplantation, islet vasculature is chimeric and is composed of host and donor endothelial cells, both of which contribute to functional blood vessels (65,66,67). These data suggest that factors that stimulate angiogenesis in isolated islets during culture and transplantation might have a positive impact on the success of the transplantation process.

Considerable effort is being devoted to discover the effects of proangiogenic factors on the success of revascularization and survival of newly transplanted islets. For example, Vegf-a overexpression in transplanted mouse islets has been shown to cause increased vascularization of and increased blood flow to newly transplanted islets, resulting in increased islet insulin content, improved recipient glucose tolerance, and increased β-cell survival (68,69,70). Similarly, overexpression of the proangiogenic growth factor angiopoeitin-1 (Ang-1), which is normally produced by islets (30), in transplanted islets leads to increased glucose tolerance, glucose-stimulated insulin secretion, islet vascular density, and islet survival (71). Interestingly, Ang-1 can stimulate pericyte migration to endothelial cells, so these studies may suggest a role for pericytes in newly transplanted islets and revascularization. Pericytes are well-described to influence endothelial cell maturation and proliferation, so proper pericyte coverage could be important in this regard. Conversely, blockade of the angiogenesis inhibitor thrombospondin-1 in transplanted islets leads to increased vascularization and improved glucose-stimulated insulin secretion (72). Prolactin overexpression in newly transplanted islets has similar beneficial effects on islet vascularization and functionality (73). Finally, coculture of human endothelial cells and mesenchymal stem cells (MSCs) with isolated islets increases islet angiogenesis and suggests a potential method for increasing vascularization of newly transplanted islets (74). Each of these studies demonstrates the utility of promoting efficient islet vascularization after transplantation and demonstrates beneficial effects on β-cell survival and function. Therefore, the elucidation of additional factors that are critical for islet vascularization may contribute greatly to the treatment of diabetes.

One fascinating new model for studying the revascularization of newly transplanted islets was recently described. Isolated islets were transplanted onto the retina of nude mice, and this procedure was able to efficiently normalize streptozotocin-induced hyperglycemia (75). In contrast to other sites of transplant, retinal islet transplant allows for real-time in vivo monitoring of islet revascularization and could provide a useful model system for future studies of islet vascularization during transplantation.

IV. Peripheral Vasculature and Insulin Delivery

A. Introduction to peripheral vasculature and insulin delivery

Insulin signaling in skeletal muscle is critical in glucose disposal, accounting for almost 90% of whole-body glucose disposal in humans (76). This is compared with adipose tissue, which is estimated to account for less than 1% of whole-body glucose disposal in humans and rodents (77,78). The role of the muscle vasculature is beginning to be appreciated as a factor that influences the myocyte’s response to insulin. Loss of skeletal muscle capillary density is observed in both insulin resistance and in type 2 diabetes, and insulin action is positively correlated with capillary density (79). In cultured myocytes, insulin activates insulin receptor and insulin receptor substrate (IRS) proteins and stimulates glucose uptake in a matter of minutes (80,81,82). This suggests that when insulin is present at the myocyte cell surface, it is able to act almost instantaneously. Although it is tempting to assume that insulin can act as swiftly in vivo, results from several groups suggest that this is not the case. Measurements of insulin in lymph, which is derived from interstitial fluid, suggest that lymph insulin concentrations are reduced for up to 3 h during an insulin infusion compared with plasma insulin levels (83). Additionally, muscle glucose utilization correlates much more strongly with lymph than with plasma insulin levels (83). These data suggest that there is a delay between the appearance of insulin in the bloodstream and its appearance in the muscle interstitium. As is the case in cultured myocytes, insulin acts very quickly upon appearance in the interstitium. In support of this latter statement, direct injection of insulin into the muscle interstitium triggers muscle glucose uptake within minutes and circumvents the delay associated with iv insulin delivery (84).

Recently, the delivery of insulin to myocytes has come under increased scrutiny for many of the reasons mentioned above. Wang et al. (85) demonstrated that after 10 min of fluorescein isothiocyanate (FITC)-insulin infusion, the majority of FITC-insulin in muscle is localized within endothelial cells, suggesting that insulin is rapidly transported from the bloodstream into endothelial cells. Interestingly, after 1 h, most FITC-insulin is still concentrated in muscle endothelial cells, suggesting that transport from endothelial cells to the muscle interstitium is a slow process (85). Several studies suggest that insulin is only degraded at low levels by endothelial cells and that muscle lymph flow is relatively slow compared with other tissues, suggesting that any clearance of insulin from the muscle interstitium is likely due to uptake by myocytes (86,87,88). These results suggest that the transendothelial transport of insulin is a rate-limiting step in muscle insulin action. In support of this conclusion, direct injection of insulin into muscle lymph causes very rapid insulin action, suggesting that direct injection sidesteps the rate-limiting step (84). Interestingly, although the interstitial passage of insulin does not appear to be a rate-limiting step under normal conditions, injection of insulin into lymph of ob/ob mice shows a delayed action as compared with lean mice, suggesting that the movement of insulin within the muscle interstitium might be impaired under conditions of insulin resistance (89).

The heterogeneity of the body’s vascular beds provides further support for a muscle-specific delay in insulin action. As mentioned above, the liver vasculature is highly permeable (5), and in fact, insulin-stimulated inhibition of hepatic glucose output occurs much more quickly than insulin-stimulated muscle glucose disposal (90). Although the more rapid effect of insulin on liver has not been directly linked to increased delivery of insulin, the higher permeability of liver vs. muscle blood vessels suggests that this is a likely cause. Interestingly, inhibition of hepatic glucose output occurs at even low levels of insulin infusion, suggesting that the highly permeable liver vasculature does not impede the transport of insulin as is the case in muscle (91). In support of this, the concentration of lymph insulin required to stimulate muscle glucose uptake is similar to the plasma insulin concentration required to inhibit hepatic glucose output (3,91).

B. Transendothelial transport of insulin

The mechanism for insulin transport across the endothelium is not clear. Several in vitro studies suggest that the endothelial uptake of insulin is mediated by the insulin receptor (86,92,93). However, the study of a transendothelial transport process in vitro is complicated by the use of cultured cells or the severing of native blood vessels. Therefore, the in vivo setting seems to be a more reliable place to study the vasculature. Indeed, in vivo results suggest that the transport of insulin across the endothelium does not appear to be saturable, even at pharmacological concentrations of insulin, suggesting that it is not a receptor-mediated process (94,95). At a minimum, these data seem to suggest that at least at high levels of insulin, transport can occur by a non-insulin receptor-mediated transcellular or paracellular process. In some respects, a paracellular transport process would be somewhat surprising given the tightness of muscle endothelium (5,6,12).

Recent research has focused on possible mechanisms for insulin transport across endothelial cells. In vitro studies in cultured endothelial cells support an insulin receptor-mediated pathway for insulin uptake because blockade of several insulin-signaling pathways inhibits this process (96). Confocal imaging studies suggest colocalization of FITC-insulin with the insulin receptor in muscle endothelial cells (85). These proteins also colocalize with caveolin-1, a protein involved in the formation of caveolae. Interestingly, caveolae are also increased in β-cell-specific VEGF-A knockouts, which display a dramatic loss of islet endothelial fenestrae, as mentioned in Section III. C. In this situation, caveolae are thought to mediate the transendothelial transport of insulin from β-cells to the bloodstream to compensate for the loss of endothelial cell fenestrae, so there is some precedent for caveolae-mediated transport of insulin across endothelial cells. Although the cellular colocalization of labeled insulin, insulin receptor, and caveloae is intriguing, further studies will be necessary to provide a conclusive link between transendothelial insulin transport in muscle and caveolae. Interestingly, transendothelial transport of insulin has been directly demonstrated by using 125I-labeled insulin in heart muscle (97).

C. Effects of insulin on blood flow

Insulin has been demonstrated to cause a number of direct effects on the vasculature. Although somewhat controversial, insulin appears to increase total blood flow within skeletal muscle. Baron (98) was the first to theorize that the ability of insulin to increase limb/muscle blood flow might be a critical part of its delivery and its ability to stimulate glucose uptake. Although this has been supported by a number of studies in both normal and obese or insulin-resistant situations (99,100,101,102,103,104,105,106,107,108,109,110,111), there are also a number of studies that failed to show an insulin-induced increase in blood flow, especially at physiological levels of insulin (112,113,114,115,116,117,118,119). This has led to some disagreement in the field as to whether insulin can increase blood flow at physiological levels (120,121). There are a number of theories to explain why an insulin-stimulated increase in blood flow is not always observed. Barrett et al. (3) argue that pharmacological doses of insulin clearly act to increase limb blood flow, but the necessity for large doses of insulin and the long durations required to observe an effect cast doubt on the normal physiological relevance of this effect. Clark (4) and Barrett et al. (3) both suggest that although there is not always an observed increase in total blood flow, insulin can influence which vessels in muscle are perfused without changing total blood flow. Specifically, this switch involves the ability of insulin to increase vasomotion, the relaxation of terminal arterioles, and shift blood flow from a nonnutritive pathway, i.e., one that has little or no exchange with the interstitial fluid surrounding the muscle fibers, to a nutritive pathway, i.e., one where blood flows through capillaries that allow for nutrient exchange with the muscle interstitium that is underperfused in the basal state (4). This insulin-induced shift provides a greater surface area for nutrient exchange and may be the primary vascular effect caused by insulin (4).

D. Insulin-induced capillary recruitment

The ability of insulin to stimulate relaxation of terminal arteries and promote flow through the nutritive pathway is termed capillary or microvascular recruitment and, in contrast to insulin-induced effects on total blood flow, seems to be gaining general acceptance (3,4). In normal muscle tissue, only about two thirds of the vasculature is normally perfused with erythrocytes at any given time (122), although other studies have demonstrated by live imaging that nearly all capillaries in the spinotrapezius muscle and diaphragm are perfused at rest (123). Potentially, differences in flow rates and the extent of perfusion between different muscle capillary beds could explain this apparent inconsistency. By shifting blood flow from nonnutritive vessels to nutritive capillaries, insulin is able to increase the available surface area for nutrient exchange and theoretically increase the surface area for its own delivery and that of glucose to myocytes (3) (Fig. 11,, A and B; and a color version in Supplemental Fig. 1, published as supplemental data on The Endocrine Society’s Journals Online web site at http://edrv.endojournals.org).

Figure 1
Transendothelial transport of insulin and glucose to muscle interstitium. A, In the normal state, insulin and glucose must travel from the blood stream across endothelial cells and potentially pericytes to reach the muscle interstitium and activate insulin ...

Two notable experimental advances greatly aided the elucidation of insulin’s ability to promote capillary recruitment: 1-methylxanthine (1-MX) metabolism (124), and contrast-enhanced ultrasound (CEU) (125). In short, 1-MX is a substrate for the endothelial cell enzyme xanthine oxidase, which is found only in smaller arterioles and capillaries and catalyzes the conversion of 1-MX to 1-methylurate (124). Disappearance of 1-MX from blood suggests an increase in available endothelial cell area that allows for 1-MX entry (124). Insulin administration significantly increases 1-MX metabolism, suggesting a dilation-mediated increase in endothelial cell area (124). CEU uses gas-filled microbubbles as a contrast agent and a surrogate for erythrocytes to measure blood flow and has demonstrated insulin-induced increases of muscle capillary volume without changes in total blood flow (125). Importantly, both 1-MX metabolism and CEU have demonstrated the ability of physiological levels of insulin to induce capillary recruitment (124,125,126,127). Insulin-induced capillary recruitment has also been investigated in skin, which has the advantage of being much easier to access and monitor (3,4,128). These results are generally consistent with those in muscle and have been reviewed in detail elsewhere (3,4).

E. Molecular mechanism of capillary recruitment

Insulin stimulates its vasodilatory actions associated with capillary recruitment through up-regulation of endothelial nitric oxide synthase (eNOS) in muscle endothelial cells (129,130,131,132) (Fig. 2A2A,, and a color version in Supplemental Fig. 2). eNOS production is induced by insulin receptor activation of the phosphatidylinositol-3-kinase (PI3K) signaling cascade, which ultimately increases eNOS expression and subsequent nitric oxide (NO) production for vasodilation (129,131,132,133). Somewhat paradoxically, insulin can also stimulate vasoconstriction by activating endothelin via the ERK 1/2 pathway (131,134) (Fig. 2B2B).). Therefore, it seems that in the normal case, vessel relaxation and constriction are tightly controlled by an organism’s metabolic state, and this suggests that perturbations to this balance might affect delivery of nutrients and insulin itself to myocytes. Indeed, vessels from obese ZF rats display reduced levels of eNOS protein compared with lean rats, and insulin treatment stimulates only vasoconstriction, which presumably contributes to the development of insulin resistance (135). Similarly, blockade of endothelin-1 receptors in obese humans results in significant vasodilation, but a similar effect is not seen in lean humans (136). A shift toward insulin-induced vasoconstriction is also observed by pharmacologically inhibiting eNOS with NO inhibitors, confirming the NO independence of this vasoconstriction (134,135). As might be predicted, eNOS knockout mice display insulin resistance (137). These data suggest that under conditions of insulin resistance, the balance of insulin action shifts toward vasoconstriction. This further exacerbates insulin resistance by reducing the access of insulin and nutrients to myocytes by decreasing nutritive flow and available capillary surface area.

Figure 2
Insulin-induced vasodilation and vasoconstriction signaling in endothelial cells and pericytes. A, Insulin signaling in endothelial cells results in increased NO production and secretion, which stimulates the dephosphorylation of myosin in pericytes and ...

F. Insulin resistance and muscle vasculature

As is the case in pancreatic islets, muscle vascular defects worsen with insulin resistance and type 2 diabetes. Numerous studies demonstrate that the insulin-induced increase in capillary recruitment is blunted in cases of insulin resistance and obesity (Fig. 1C1C).). For example, insulin-resistant ZF rats show reduced capillary recruitment (138). Similarly, insulin resistance induced by various agents such as TNF-α, intralipid/heparin, glucosamine, or α-methylserotonin also reduces capillary recruitment (139,140,141,142). Importantly, the extent of muscle capillary recruitment is positively correlated with glucose uptake (140,141), suggesting a possible causal relationship between these two processes. Human type 2 diabetic patients demonstrate a reduced muscle capillary permeability-surface area in response to hyperinsulinemia, suggesting a defect in insulin-induced capillary recruitment (143). A likely mechanism for these observations, as mentioned in Section IV. E, is the insulin resistance-induced decrease of eNOS activation, which leads to reduced vasodilation. These data suggest that muscle vascular dysfunction accompanies insulin resistance.

Whether insulin resistance causes vascular dysfunction or vice versa is not entirely clear. Evidence suggests that muscle vascular defects are among the earliest phenotypes observed as insulin resistance progresses and that diminished NO production and increased vasoconstriction precede the development of type 2 diabetes (144). Due to the importance of insulin signaling in capillary recruitment, it would not be surprising if reduced eNOS activation was mediated by the same mechanism as desensitization of myocyte insulin signaling. Insulin resistance in myocytes has been demonstrated to occur downstream of insulin receptor binding, likely at the level of the IRS proteins (145,146). One can imagine that the same desensitization of insulin signaling could blunt the PI3K pathway in endothelial cells and subsequent eNOS activation, thus leading to reduced vascular dilation. The subsequent reduction in available capillary surface area would likely reduce transendothelial insulin transport to myocytes and further blunt myocyte insulin signaling, continuing a vicious cycle. As the potential involvement of the insulin receptor in transendothelial transport is further elucidated, defects in this process might also become important for the development of insulin resistance.

Several animal models are informative regarding the effects of insulin on muscle vasculature. Prominent among these models are the vascular endothelial cell insulin receptor knockout (VENIRKO) mice, which somewhat surprisingly, are not insulin resistant (147). An initial reaction to this result might be that insulin-mediated vasodilation is not a critical process for muscle glucose disposal. This suggests that insulin delivery to the periphery is possible without the endothelial cell insulin receptor under normal circumstances. However, both eNOS and endothelin-1 mRNA levels are reduced in the VENIRKO mice (147), suggesting that the lack of insulin resistance in these mice may not be surprising because the signals for both vasoconstriction and vasodilation are reduced, resulting in no net changes in vasomotion. When considering these results, it is important to note that in lean mice, creating insulin resistance has historically been somewhat complicated because even muscle-specific knockout of the insulin receptor (MIRKO mice), one of the most critical proteins involved in muscle insulin signaling, results in normal glucose tolerance and insulin sensitivity (148). Finally, the insulin-like growth factor-1 receptor, which is found at high levels in muscle endothelial cells (149), could also compensate for loss of the insulin receptor. Additionally, as mentioned in Section IV. E, eNOS knockout mice are also insulin resistant (137). No studies linking knockout of endothelin-1 to amelioration of insulin resistance have been reported, but this result might be predicted.

G. Exercise-induced vascular changes

Exercise has been demonstrated to promote many of the same changes in the muscle vasculature as insulin (150). Exercise efficiently increases muscle capillary recruitment, as well as total muscle blood flow (151). Interestingly, the mechanism for insulin- and exercise-induced increases in muscle perfusion appear distinct because the exercise mechanism is NO-independent (152). Exercise-induced vascular changes are not blunted by insulin resistance or obesity (153). These data suggest that even in cases of prolonged disease progression, exercise might be able to remedy some of the effects of insulin resistance on the muscle vasculature.

V. Vascular Pericytes: More Than Inert Contractile Cells

A. Introduction to pericytes

Although pericytes were first described almost 150 yr ago, they are still not entirely understood. Charles Rouget was the first to identify pericytes in 1873 when he described a population of perivascular cells that he regarded as contractile elements (154). In 1923, Zimmermann named these cells “Rouget cells” after their discoverer, or “pericytes” because of their location in proximity to endothelial cells (peri, around; cyte, cell) (155). Together, pericytes and the related vSMCs make up a class of cells, termed mural cells, which provide support to blood vessels of all sizes (156). By convention, the mural cells associated with larger vessels such as arteries and veins are called vSMCs, whereas those associated with smaller vessels, such as capillaries, arterioles, and venules, are termed pericytes (157). vSMCs are highly contractile, typically express high levels of α-smooth muscle actin (α-SMA), and can form multiple concentric layers around blood vessels (156). Additionally, vSMCs have their own basement membrane, which is rich in elastin and fibrillar collagen and is separate from the vascular basement membrane (156,158). Pericytes, on the other hand, typically form a single discontinuous layer around smaller vessels (157). They are intimately associated with endothelial cells and reside in a shared basement membrane that is produced by both pericytes and endothelial cells (159). Pericytes typically express high levels of cell-surface chondroitin sulfate proteoglycan neuron-glial 2 (NG-2), which is commonly used as a histological pericyte marker (155).

Pericyte coverage and morphology can vary between different vascular beds. Pericytes in certain tissues have acquired specialized names, such as Ito cells in liver (named after their discoverer, Toshio Ito) and mesangial cells in the kidney glomerulus (155,160). The highest level of pericyte coverage is seen in the central nervous system, where pericytes are thought to play an important role in the formation and maintenance of the blood-brain barrier (161,162).

Although pericytes were originally thought to be solely involved in contractile processes, they have recently been shown to have many additional functions. Pericytes, like vSMCs, can indeed mediate vessel contraction and influence vascular diameter and capillary blood flow (163). Loss of pericytes leads to excessive endothelial membrane folding and luminal cytoplasmic protrusions, which likely impacts vessel perfusion and possibly nutrient exchange (164). Pericytes can communicate with endothelial cells through a variety of signaling pathways, and they promote endothelial cell differentiation and maturation (165). Pericytes are also thought to limit endothelial cell proliferation; thus, loss of pericytes can lead to endothelial cell hyperplasia (164). Pericytes also play a critical role in angiogenesis, because they are typically found near the tips of sprouting vessels and where they are thought to guide the sprouting processes by expressing VEGF (166,167,168,169,170).

B. Platelet-derived growth factor-B: a key mediator of pericyte function

Platelet-derived growth factor B (PDGF-B) and its receptor, PDGF receptor-β (PDGFRβ), are critically involved in pericyte recruitment and proliferation (157). PDGF-B belongs to the PDGF family of proteins, which shares structural homology with the VEGF protein family (171,172). In addition to PDGF-B, there are three other mammalian PDGF family members, PDGF-A, PDGF-C, and PDGF-D (172). PDGF was originally identified as a constituent of whole blood that was absent in cell-free plasma (173,174,175) and was later purified from human platelets (176,177,178,179). The PDGF family members function as homo- and heterodimers (180). PDGF-B is normally expressed in vascular endothelial cells, megakaryocytes, and neurons, whereas PDGF-A and PDGF-C are highly expressed in epithelial cells, muscle, and neuronal progenitor cells (172).

There are two PDGF receptors, PDGFRα and PDGFRβ. Ligand binding causes homo- or heterodimerization of PDGFRs, which activates their cytoplasmic tyrosine kinase domains and results in autophosphorylation of several tyrosine residues in their cytoplasmic tails (181). The phospho-tyrosine residues create docking sites for a variety of adaptor proteins, leading to activation of a wide variety of signaling pathways, including the RAS-MAPK, JNK/SAPK, PI3K/AKT, and protein kinase C (PKC) pathways (182,183). Interestingly, PDGFR signaling through different pathways appears to be additive rather than producing distinct outcomes. In a tour de force study, Tallquist et al. (184) created an allelic series of tyrosine to phenylalanine mutations, which resulted in blunted but qualitatively similar developmental effects. Ligand occupancy promotes internalization and subsequent lysosomal degradation of PDGFR complexes, limiting the duration of signaling (185,186,187).

PDGFRα binds both PDGF-A and PDGF-B with high affinity, whereas PDGFRβ binds only PDGF-B with high affinity (180). However, the receptor complex thought to mediate in vivo PDGF-B signaling is the PDGFRβ homodimer (157,158,172). In addition, outside of human platelets, the expression patterns of PDGF-A and PDGF-B are generally nonoverlapping, suggesting that the PDGF-AB heterodimer, which can bind to both PDGFRs, might have limited in vivo significance (180,183).

PDGF-A and PDGF-B are the best described PDGF ligands. Although PDGF-A is involved in a variety of diverse developmental and organogenesis processes (172), the major function of PDGF-B is its role in stimulating the migration of pericytes and vSMCs to growing blood vessels (172). PDGF-B is secreted as a homodimer from endothelial cells, where it is retained on the cell surface by a C-terminal heparan-sulfate proteoglycan-binding retention motif (188,189). PDGFRβ on the surface of pericytes and vSMCs binds to PDGF-B, thus recruiting mural cells to the endothelial cell wall. Loss of the retention motif leads to decreased retention of PDGF-B on the cell surface (188,189,190). Lindblom et al. (191) have generated and characterized mice in which the PDGF-B retention motif has been deleted by targeted mutagenesis, causing a reduction in PDGF-B signaling. These mice display a loss of pericyte density, pericyte detachment, and abnormal capillary morphology in the developing brain, kidney, and retina, as well as the postnatal kidney and retina (191). PDGF-B and PDGFRβ knockout mice display similar, but more severe defects: a drastic loss of pericytes, widespread vascular leakage, general heart defects, and perinatal lethality (164,192,193,194,195).

Although PDGF-B and PDGFRβ are critically important for pericyte recruitment, the presence of pericytes in some organs of PDGF-B/PDGFRβ knockouts suggests that other factors are also important for pericyte biology. Notably, TGF-β is an important mediator of vSMC/pericyte differentiation (196,197). Additionally, Ang-1 and its receptor, endothelium-specific receptor tyrosine kinase-2, are important for signaling from pericytes/vSMCs to endothelial cells and play a role in pericyte recruitment (157).

Although the importance of PDGF-B signaling and pericyte coverage in the microvasculature has been well described, little has been reported regarding any influence pericytes might have on insulin secretion and peripheral insulin action. Pericytes are found at relatively high densities in skeletal and cardiac muscle (198,199) and in pancreatic islets (200). PDGF-B signaling can promote β-cell proliferation in vitro, but only under conditions of PDGFRβ ectopic overexpression; endogenous PDGFRβ expression in β-cells is quite low (201). Therefore, PDGF-B does not directly signal to β-cells.

C. Diabetic complications: a key role for pericytes

Pericytes play a central role in diabetic complications. Loss of pericytes is one of the first observable changes in diabetic retinopathy and is ultimately followed by increased vascular permeability (202). Interestingly, Geraldes et al. (203) recently demonstrated that glucose increases the expression of PKC-δ, which results in down-regulation of PDGFRβ/AKT survival signals and pericyte death. The vasculature also plays a clear role in the progression of nephropathy (204). Knockout of PDGF-B/PDGFRβ or loss of PDGF-B retention results in defects in the pericyte-like mesangial cells, leading to defective glomerulogenesis, glomerulosclerosis (191,194,195), and proteinurea (191), a hallmark of diabetic nephropathy. Diabetic neuropathy is characterized by reduced blood flow, capillary basement membrane thickening, and pericyte degeneration (205). Tilton et al. (206) used transmission electron microscopy to study pericyte morphology and density in a variety of skeletal muscles from nondiabetic and diabetic humans. Similar to what is observed in diabetic retinopathy, they noted an increased degeneration of pericytes in the type 2 diabetic muscles. Pericyte changes are therefore associated with diabetes and are found in most of the microvascular complications associated with diabetes.

D. Are pericytes multipotent progenitor cells?

One exciting new field of pericytes involves their potential role as MSC-like progenitor cells. MSCs, also known as multipotent mesenchymal stromal cells, are undifferentiated, self-renewable cells that are present in bone marrow and mesenchymal tissues (207). Interest in MSCs in relation to diabetes intensified when it was reported that transplanted bone marrow MSCs initiated pancreas regeneration and improved diabetes in mice and humans (208,209,210). However, other studies did not demonstrate any evidence for transdifferentiation of bone marrow cells into β-cells (211,212), so this process could be dependent on the stage of diabetes progression and/or the isolation and transplantation techniques. Transplantation of purified MSCs was likewise able to normalize hyperglycemia and promote islet growth in streptozotocin-treated mice (213). Crisan et al. (214) recently demonstrated that pericytes isolated from human mesenchymal tissues, including skeletal muscle, pancreas, and adipose tissue, were able to serve as multilineage progenitor cells reminiscent of MSCs. Specifically, this study demonstrated that purified pericytes from any of these organs, when cultured in a tissue-specific growth medium, could differentiate into myocytes, adipocytes, osteocytes, and chondrocytes (214). Although pericytes isolated from pancreas were used as progenitor cells, the authors did not report on the ability of isolated pericytes to differentiate into β-cells or any other pancreatic cell type. This presents the intriguing question: can pericytes serve as β-cell progenitors? In vivo lineage-tracing experiments using a tetracycline-inducible CRE recombinase under the control of the adipogenic peroxisome proliferator-activated receptor-γ promoter to indelibly mark cells with β-galactosidase demonstrate that adipocyte progenitors are perivascular cells that express several pericyte markers, including NG-2, PDGFRβ, and α-SMA (215). One can imagine using a similar system with a β-cell-specific CRE recombinase such as Pdx1-CRE to determine whether islet pericytes can analogously serve as progenitors to β-cells.

E. Pericytes in normal islet function

The role of pericytes in normal islet function is not completely understood, but islet pericyte changes associated with a number of pathological conditions have been described. In obese animals, islet pericytes become more hypertrophied and assume vSMC characteristics (46). Notably, pericytes take on more of a smooth muscle cell-like appearance, and it has been speculated that this is potentially in response to increased islet blood pressure (46). We and others have similarly observed an increase in α-SMA and NG-2 staining densities in islets from ob/ob mice, consistent with obesity-induced pericyte hypertrophy (Ref. 216 and Supplemental Fig. 3). Similarly, hypertensive Ren2 rat islets have increased pericyte proliferation, migration, hypertrophy, and α-SMA staining (217). In rats overexpressing human islet amyloid polypeptide, there is a reduction in both β-cell mass and islet capillary density, along with increased pericyte apoptosis and loss (218). Finally, in a rat model of type 2 diabetes, the matrix between islets and the surrounding exocrine tissue widens, due to an increase in pericytes and inflammatory cells in this region (219). The authors of these studies noted evidence of pericyte differentiation into stellate cells, but the use of α-SMA as a stellate-cell marker seems somewhat questionable due to its well-described role as a vSMC marker (155).

F. Pericytes in islet tumors

The role of pericytes in islet-cell tumors has been intensively investigated (220,221,222,223,224,225,226,227). PDGFRs are expressed in tumor pericytes, and treatment with a drug that selectively inhibits PDGFRα and PDGFRβ blocks further growth of end-stage tumors by causing pericyte detachment and disrupting tumor vasculature (220). Pericyte loss induced by treatment with imatinib mesylate improves the efficacy of metronomic chemotherapy by rendering endothelial cells more sensitive to the actions of the cytotoxic drugs (224). Treatment with imatinib mesylate, metronomic chemotherapy, and a selective VEGFR inhibitor elicits regression of solid islet tumors and increases median survival (224). Some of the effects of PDGFRβ inhibition in islet tumors are mediated by elimination of PDGFRβ-positive perivascular progenitor cells, which can differentiate into mature pericytes (226). These data suggest that lack of pericytes improves drug delivery to tumor cells and might be beneficial for treatment.

Although the absence of tumor pericytes might be beneficial for drug delivery, it is also associated with increased tumor metastasis. β-Cell tumors that are deficient for neural cell adhesion molecule have leaky blood vessels with detached pericytes, which correlates with an increased incidence of metastasis (225). In agreement with this, PDGF-B retention-deficient mice with β-cell tumors exhibit increased metastasis, demonstrating a direct causal link between defective pericyte recruitment and increased metastasis (225). These findings directly translate to humans, because decreased α-SMA-positive pericyte coverage of tumor vessels correlates with increased metastasis and results in a poorer prognosis (228). Thus, although inhibition of PDGFR is an attractive option for improving tumor treatment, the benefits must be weighed against the increased metastatic potential associated with reduced tumor pericyte coverage.

G. A role for PDGF-B signaling in glucose uptake?

PDGFRβ has been investigated in relation to insulin signaling because it can activate several of the same signaling pathways as the insulin receptor. Specifically, the ability of PDGFRβ to activate AKT/PI3K signaling (183) has been investigated in relation to glucose uptake into myocytes and adipocytes. PDGF-B signaling can induce glucose transporter type 4 translocation in both cultured adipocytes (229) and mouse skeletal muscle (230). However, due to minimal endogenous expression of the receptor, this is only possible under conditions involving overexpression of PDGFRβ. These actions of PDGF-B are mediated independently of IRS-1 activation (229,230). Interestingly, exogenous overexpression of PDGFRβ in skeletal muscle has been used to demonstrate that defects in the insulin signaling pathway independent of IRS-1 can lead to insulin resistance (231). These data suggest that any effect of PDGF-B on insulin action is unlikely due to a direct effect of PDGF-B on parenchymal cells.

Although PDGF-B does not normally signal directly to myocytes, we have recently demonstrated that loss of PDGF-B activity does impact peripheral insulin sensitivity (200). In ob/ob mice, loss of PDGF-B retention causes decreased in vivo insulin secretion without a change in glucose tolerance (200). These mice have defective pericyte coverage in peripheral tissues involved in insulin action (200). Loss of pericytes can lead to increased vascular leakage (164,191,192,193,194,195), and indeed these mice display increased vascular permeability, especially in heart (200). Ultimately, this leads to increased transendothelial transport of insulin and increased whole-body insulin sensitivity (200). These data demonstrate an important and novel role for pericytes and PDGF-B in delivery of insulin to peripheral tissues.

H. Inhibition of PDGFRβ and diabetes therapy

One potentially interesting link between PDGF-B activity and diabetes concerns the effect of imatinib mesylate (Gleevec). Imatinib mesylate inhibits several receptor tyrosine kinases, including c-abl, c-kit, and PDGFRβ (232). Several studies have reported that imatinib mesylate lowers fasting blood glucose levels in diabetic patients treated for chronic myeloid leukemia (233,234,235). Another study reported no effect of imatinib mesylate treatment on glucose levels (236). However, Han et al. (237) recently demonstrated that treatment of db/db mice with imatinib mesylate drastically improves peripheral insulin sensitivity. The authors observed improved insulin sensitivity and decreased in vivo insulin secretion in response to a glucose challenge, but they attributed these effects to an amelioration of c-abl-induced endoplasmic reticulum stress in liver and adipose tissues. However, it is also possible that the observed increase in insulin sensitivity may also involve PDGFRβ inhibition. In accordance with this, treatment of mice with imatinib mesylate or a soluble form of PDGFRβ both prevents and reverses type 1 diabetes, whereas treatment with a c-kit inhibitor had little effect (238). None of these studies investigated the effect of imatinib mesylate treatment on pericyte coverage of islets or peripheral tissues, which could provide a potential mechanism for the improvement of diabetes. In support of this possibility, tumor-bearing mice treated with imatinib mesylate demonstrate a decrease in pericyte coverage and increased vessel leakiness, suggesting that a similar improvement of vascular permeability in peripheral tissues could improve insulin or nutrient delivery and possibly explain the effects of imatinib mesylate on diabetes (239). This suggests that further investigation into the effects of imatinib mesylate and other PDGFRβ inhibitors on diabetes treatment and pericyte coverage may be informative.

I. A role for pericytes in insulin-induced hemodynamic changes

In addition to the role for pericytes in insulin delivery to myocytes, pericytes and vSMCs also play an important role in insulin-induced capillary recruitment. NO produced by endothelial-expressed eNOS diffuses into vSMCs where it binds to and activates the heme moiety of guanylyl cyclase (240,241) (Fig. 2A2A).). This results in an increase of the local concentration of cGMP, which increases cGMP-dependent protein kinase G (PKG) signaling (240,241). This results in activation of myosin light-chain phosphatase and the opening of KATP channels (240). NO can also directly nitrosylate KATP channels, resulting in hyperpolarization of the vSMC plasma membrane and inhibition of calcium entry (240). Each of these NO-mediated effects results in vSMC relaxation and concomitant vasodilation. Conversely, endothelin-1 signals through its G protein-coupled receptors on the surface of vSMCs, ETA and ETB, to activate PKC signaling that results in increased calcium levels and vascular contraction (242,243,244) (Fig. 2B2B).). Interestingly, PKC signaling can be activated by lipid by-products like diacylglycerol and long-chain acyl-coenzyme A, which are increased in obesity (145) and can further promote vasoconstriction and exacerbate insulin resistance. Endothelin-1 and NO mediate their vasoconstriction and vasodilation effects on pericytes through similar mechanisms (163).

Modulation of vSMC or pericyte coverage and its effects on insulin-induced vascular constriction and dilation have not been directly studied, although one can imagine that loss of vSMCs or pericytes could reduce the ability of vessels to respond to insulin-induced changes in vascular tone. Additionally, loss of vSMCs/pericytes might lead to general increases in vessel dilation and muscle perfusion, resulting in increased capillary surface area and insulin/nutrient transport to muscle interstitium. Conversely, vSMC/pericyte hyperplasia might result in increased basal vascular contraction. Notably, vSMCs isolated from ZF rats showed reduced PKG activation by NO and cGMP, suggesting that insulin resistance also impedes this step in insulin-induced vasodilation (245). This defect was thought to be due to increased levels of superoxide anions because it was rescued by antioxidant treatment (245). In keeping with this, NO can be consumed by reactive oxygen species, resulting in the production of peroxynitrite (246). This suggests that increased metabolism and increased generation of metabolic by-products like reactive oxygen species could begin to explain a mechanism for defective eNOS/NO signaling in insulin-resistant animals and humans.

PDGF-B could play a direct role in controlling vascular dilation and constriction. Like insulin, PDGF-AB and to a lesser extent PDGF-A and PDGF-B increase eNOS expression (247). This suggests that, in addition to decreasing pericyte coverage, defects in PDGF signaling could directly contribute to the development of insulin resistance by reducing eNOS levels and increasing vasconstriction.

VI. Summary/Conclusions

Type 2 diabetes is a growing worldwide epidemic. The involvement of the vasculature in the processes of islet development, insulin secretion, and peripheral insulin action is undeniable. Although most recent research has focused on endothelial cells, the vascular pericyte is an intriguing candidate to play a role in these processes. From their involvement in diabetic complications to their function as mediators of insulin-induced vasodilation and vasoconstriction, the available information suggests a role for pericytes in the development of insulin resistance and type 2 diabetes. Future research will be critical to elucidate the role of the vascular pericyte in these processes.

Supplementary Material

Supplemental Data:

Acknowledgments

We are grateful to Robin Davies and Laura Vanderploeg for expert assistance in preparing Figs. 11 and 22.. We thank William Dove, Jon Odorico, Anath Shalev, and Xin Sun for critical review of the manuscript and helpful discussion.

Footnotes

This work was supported by National Institute of Diabetes and Digestive and Kidney Diseases Grants DK66369 and DK58037, National Institutes of Health Training Grant T32GN07215 (to O.C.R. and S.M.R.), and a Wisconsin Alumni Research Foundation fellowship (to O.C.R.).

Disclosure Summary: The authors have nothing to disclose.

First Published Online February 17, 2010

Abbreviations: Ang-1, Angiopoeitin-1; CEU, contrast-enhanced ultrasound; eNOS, endothelial NO synthase; FITC, fluorescein isothiocyanate; HIF-1α, hypoxia-inducible factor-1α; IRS, insulin receptor substrate; MSC, mesenchymal stem cell; 1-MX, 1-methylxanthine; NG-2, neuron-glial 2; NO, nitric oxide; PDGF, platelet-derived growth factor; PDGFR, PDGF receptor; Pdx-1, pancreatic and duodenal homeobox 1; PI3K, phosphatidylinositol-3-kinase; PKC, protein kinase C; PKG, protein kinase G; RIP, rat insulin promoter; α-SMA, α-smooth muscle actin; Vegf-A, vascular endothelial growth factor-A; vSMC, vascular smooth muscle cell; ZF, Zucker fatty (rats).

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