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. Author manuscript; available in PMC: 2021 Dec 1.
Published in final edited form as: Exp Gerontol. 2020 Oct 25;142:111126. doi: 10.1016/j.exger.2020.111126

Aging differentially impacts vasodilation and angiogenesis in arteries from the white and brown adipose tissues

Torikul Islam Md 1, Grant D Henson 2, Daniel R Machin 3, R Colton Bramwell 3, Anthony J Donato 1,2,3,4,5, Lisa A Lesniewski 1,2,3,4
PMCID: PMC8407014  NIHMSID: NIHMS1732256  PMID: 33203620

Abstract

Aging adipose tissues (ATs) manifest reduced vascularity and increased hypoxia and inflammation that contribute to local and systemic metabolic dysfunction. However, the mechanisms that underlie these age-related changes are incompletely understood. In this study, we sought to examine insulin-stimulated vasodilation and angiogenesis in the arterial vasculature from three major AT depots, perigonadal white (pgWAT), subcutaneous white (scWAT) and brown (BAT) from young and old mice. Here, we demonstrate that in young mice, insulin-stimulated vasodilation is lower in feed arteries from pgWAT compared to scWAT (p<0.05), but no differences were found between feed arteries in other AT depots (p>0.05). Insulin-stimulated vasodilation was lower in old compared to young feed arteries from all three AT depots (p<0.05 for all). In the presence of endothelial nitric oxide synthase inhibitor, L-NAME, insulin-stimulated vasodilation was decreased in young (p<0.05), but was unaffected in old (p>0.05) from all AT depots. We also observed no age-related differences in endothelium-independent dilation, as assessed by sodium nitroprusside (p>0.05). We next investigated angiogenic capacity of the vasculature in these AT depots. In young mice, BAT vasculature demonstrated the highest angiogenic potential, followed by pgWAT and scWAT. We found that aging decreased angiogenic sprout formation in pgWAT and BAT (both p<0.05), but increased angiogenic potential in scWAT (p<0.05), indicating dissimilar impact of aging on angiogenesis in different AT depots. Collectively, these data suggest that aging leads to a consistent impairment in insulin-stimulated vasodilation and reduction in NO bioavailability in all three AT, although aging differentially impacts angiogenic capacity across different AT depots.

Introduction

Aging is an independent risk factor for cardiovascular diseases (CVDs) and metabolic dysfunction, both of which are intricately intertwined (Chia, Egan, & Ferrucci, 2018; Donato, Machin, & Lesniewski, 2018; Pi, Xie, & Patterson, 2018). Advancing age impairs endothelium-dependent vasodilation, leading to the development of CVD (Donato et al., 2018; Seals, Jablonski, & Donato, 2011). Furthermore, aging dysregulates organ blood flow, thus impairing the provision of nutrients and oxygen supply (Mitchell, 2008; Zhang et al., 2011). Subsequently, these defects lead to impaired organ function, local and systemic inflammation, oxidative stress, insulin resistance and other age-related diseases (McCarron et al., 2019; Mitchell, 2008; Ye, 2009; Zhang et al., 2011). Vasodilation and angiogenesis are two key mechanisms by which the vasculature adapts to changes in the demand for blood flow (Azad, Ghahremani, & Yang, 2019; Bagher & Segal, 2011; Carmeliet & Jain, 2011). Therefore, understanding age-related changes in organ specific vasodilation and angiogenesis in response to stimuli such as insulin and vascular endothelial growth factor (VEGF) are critical to the development of novel therapies for age-related metabolic dysfunction.

Adipose tissue (AT) is an active endocrine organ that plays a central role in systemic metabolic and cardiovascular homeostasis (Cinti, 2012). Different AT depots such as perigonadal white AT (pgWAT), subcutaneous white AT (scWAT) and brown AT (BAT) play independent and critical roles in energy metabolism (Cinti, 2005; Saely, Geiger, & Drexel, 2012). For example, pgWAT serves as the storage site of triglycerides and maintains lipid homeostasis (Cinti, 2012; Rosell et al., 2014). Moreover, pgWAT is a major source of metabolic hormones leptin and adiponectin, which play key roles in systemic energy metabolism (Stern, Rutkowski, & Scherer, 2016). Although less well defined in mice (Fischer, Csikasz, von Essen, Cannon, & Nedergaard, 2016); in humans, the primary role of scWAT is to maintain thermal insulation and an increase in scWAT is correlated with lower CVD risk (Porter et al., 2009). BAT generates heat via mitochondrial uncoupling and thus plays an important role in the maintenance of body temperature in rodents as well as in energy expenditure (Moonen, Nascimento, & van Marken Lichtenbelt, 2019; Saely et al., 2012). Collectively, the AT depots are critical regulators of metabolic function and thus may serve as a target to ameliorate age-related metabolic dysfunction.

Advancing age is accompanied by diminished AT vascularity and blood flow, immune cell infiltration and inflammation, and insulin resistance (Palmer & Kirkland, 2016). However, the underlying mechanisms that contribute to these aforementioned processes are unclear. We have previously demonstrated that advancing age results in impaired acetylcholine-stimulated vasodilation in pgWAT feed arteries that is associated with systemic metabolic dysfunction in mice (Donato et al., 2014). Along with the skeletal muscle, ATs are the major sites of insulin-dependent glucose and free fatty acids uptake, therefore insulin-stimulated vasodilation of the feed arteries in these tissues may be more functionally relevant than acetylcholine-stimulated vasodilation (Satoh, 2014). Although it is appreciated that insulin-stimulated vasodilation facilitates increases in blood flow to skeletal muscle that leads to increases in glucose uptake (Baron et al., 1995), little is known about insulin-stimulated vasodilation of different AT depots or the impact of aging on this vasodilation. Likewise, we have previously demonstrated that angiogenic potential of pgWAT vasculature declines with aging, likely contributing to AT dysfunction (Donato et al., 2014). However, the angiogenic potential across different AT depots or potential age-related alterations has not yet been examined.

In this study, we sought to examine insulin-stimulated vasodilation in feed arteries from three major AT depots, as well as examine the impact of advanced age on insulin-stimulated vasodilatory responses in these feed arteries. We also sought to examine the angiogenic potential of the three major AT depots and elucidate the impact of aging.

Methods

Ethical approvals.

All animal procedures conformed to the Guide for the Care and Use of Laboratory Animals: Eighth Edition (Carbone, 2012) and were approved by the Animal Care and Use Committee at the University of Utah and Salt Lake City Veteran Affairs Medical Center (SLC-VAMC).

Animals.

Young (5-7 mo) and old (27-29 mo) male B6D2F1 mice were acquired from Charles River Inc, and the NIH aging colony that is maintained at Charles River Inc. All mice were housed in standard shoe box cages in the SLC-VAMC animal care facility on a 12:12 light:dark cycle. Mice were fed a standard rodent chow and water ad libitum. Prior to tissue harvest, mice were euthanized via exsanguination by cardiac puncture while under isoflurane anesthesia (Lesniewski et al., 2017).

AT volume.

pgWAT, scWAT and BAT volume were assessed using a small animal computerized tomography (Micro-CT), as described previously (Donato et al., 2014). Briefly, scanning was performed in anaesthetized animals (2% isoflurane/O2 gas) using a Quantum FX Micro CT Scanner (Perkin-Elmer, Waltham, MA). Voltage and current were set at 50 kV and 200 μA, respectively and the images were captured over a 4.5 min interval. The abdominal region was defined superiorly by the diaphragm and inferiorly by the pelvic epiphysis. Analysis was conducted with Caliper Analyze 11.0 (Analyze Direct, Overland Park, KS). pgWAT, scWAT and BAT were segmented in the sagittal plane and volume measurements obtained with the region of interest module. AT volumes are expressed both as absolute and relative to body mass.

Ex-vivo vasodilatory function.

AT feed arteries were dissected from scWAT, pgWAT, and BAT, cleared of surrounding tissue and cannulated in the stage of pressure myograph (DMT Inc, Atlanta, GA, USA). Arteries were pre-constricted with 2 μM phenylephrine, and endothelium-dependent dilation and the contribution of nitric oxide (NO) to dilation were measured in response to the cumulative addition of insulin (0.01 to 10 nM) in the absence or presence of NO synthase inhibitor, L-NAME (0.1 mmol L−1, 30 min), as described previously (Durrant et al., 2009). Endothelial-independent dilation was assessed in response to inorganic NO donor sodium nitroprusside, SNP (1 × 10−9 to 1 × 10−4 mol L−1) (Durrant et al., 2009). Arterial diameters were measured using MyoView software (DMT, Inc., Atlanta, GA, USA). All dose response data are presented as percent of possible dilation after pre-constriction to phenylephrine. Arteries failing to achieve ≥20% preconstriction were excluded.

Angiogenic capacity.

The angiogenic capacity of adipose explants was assessed by quantifying sprout formation in vitro, as described previously (Gealekman et al., 2011). Briefly, a small (~1 mm2) explant of the pgWAT, scWAT or BAT was embedded in a collagen matrix and cultured in standard medium supplemented with vascular endothelial growth factor (VEGF: 0.5 mg/mL). Growth medium changes were performed every other day. Five days after explanting, sprouts were visualized and images were captured with phase contrast on an inverted microscope (Nikon Instruments, Melville, NY).

Statistics.

Statistical analyses were performed with SPSS software and data are presented as means ± SEM. Differences between age or fat depot were assessed by two-way ANOVA (animal characteristics, sprouting, maximal responses) or repeated measures ANOVA (concentration response curves) with LSD post hoc test where appropriate. Significance was set at p<0.05.

Results

Animal characteristics.

Body mass was higher in old (27-29 mo) compared to young (5-7 mo) mice (p<0.05) (Table 1). Heart mass, both absolute and normalized to body weight, was higher in old than young mice (both p<0.05) (Table 1). AT mass and volume of all three AT depots assessed were lower in old mice compared to young mice (all p<0.05) (Table 1). The characteristics of the feed arteries from different AT depots are summarized in Table 2. Briefly, arterial diameter was ~120-180 μm and arteries constricted ~35-55% to phenylephrine prior to functional assessments. pgWAT artery diameter was lower in old mice compared to young mice (p<0.05) and scWAT artery diameter tended to be lower in old mice compared to young mice (p=0.09). No age-related difference in BAT artery diameter was found. Preconstriction to phenylephrine was similar in arteries from each AT depot between the young and old mice (all p>0.05).

Table 1.

Animal Characteristics

Young Old P value
Body mass (g) 33.6 ± 0.6 36.7 ± 0.9 0.008
Heart mass (mg) 176 ± 8 224 ± 7 0.001
Heart mass/body mass (mg/g) 5.1 ± 0.2 6.1 ± 0.3 0.032
Adipose tissue volume (mm3)
   pgWAT 2.55 ± 0.21 1.65 ± 0.25 0.012
   scWAT 0.78 ± 0.09 0.37 ± 0.05 0.000
   BAT 0.13 ± 0.00 0.08 ± 0.00 0.000
Adipose tissue mass (mg)
   pgWAT 589 ± 47 321 ± 33 0.000
   scWAT 362 ± 34 180 ± 14 0.000
   BAT 89 ± 8 64 ± 5 0.020
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Data are shown as Mean ± SEM; N=11-13/group

Table 2.

Adipose tissue artery characteristics

Young Old P value
pgWAT Artery Diameter (μm) 179 ± 10 137 ± 13 0.010
pgWAT Artery Preconstriction (%) 47 ± 6 48 ± 8 0.600
scWAT Artery Diameter (μm) 163 ± 7 148 ± 12 0.092
scWAT Artery Preconstriction (%) 39 ± 6 43 ± 6 0.393
BAT Artery Diameter (μm) 177 ± 16 173 ± 16 0.232
BAT Artery Preconstriction (%) 45 ± 5 39 ± 4 0.600
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Data are shown as Mean ± SEM; N=11-13/group

Insulin-stimulated vasodilation and sensitivity differs between feed arteries from pgWAT, scWAT and BAT and is impaired with aging.

We first examined the insulin-stimulated vasodilation in feed arteries from different AT depots. Insulin-stimulated vasodilation in feed arteries from the pgWAT was lower compared to scWAT arteries (p<0.05), but there was no difference between scWAT and BAT feed arteries or between pgWAT and BAT feed arteries (p>0.05) (Figure 1A). Likewise, insulin-stimulated maximum dilation was lower in pgWAT (~30%) feed arteries compared to scWAT (~50%) feed arteries (p<0.05), but no difference was found between scWAT and BAT (~40%) feed arteries or between pgWAT and BAT feed arteries (p>0.05) (Figure 1B). In the presence of L-NAME, insulin-stimulated vasodilation was ~50-60% lower in all three arteries compared to insulin alone, indicating a large contribution of NO to insulin-stimulated vasodilation across all AT depots (Figure 1). To assess the sensitivity of the arteries, we calculated EC50 (the concentration that evokes 50% of the maximum response) and found that scWAT feed arteries demonstrated higher sensitivity to insulin compared to pgWAT and BAT feed arteries (scWAT: 0.75 ± 0.15; pgWAT: 2.20 ± 0.57; BAT: 1.53 ± 0.38; p<0.05). No difference was found between pgWAT and BAT feed arteries (p>0.05) (Figure 1).

Figure 1: Insulin-stimulated vasodilation in adipose tissue feed arteries from young and old mice.

Figure 1:

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(A, B) Concentration response curve and maximal vasodilation to insulin in young pgWAT, scWAT and BAT feed arteries (N=7-14, *p<0.05 vs pgWAT). (C) Concentration response curves to insulin in pgWAT feed arteries in the absence or presence of the eNOS inhibitor, L-NAME (N; young=11, old=13). (D) Maximal insulin-stimulated vasodilation of pgWAT feed arteries (N; young=11, old=13). (E) Concentration responses to insulin in scWAT feed arteries in the absence or presence of the eNOS inhibitor, L-NAME (N; young=15, old=14). (F) Maximal vasodilation to insulin in scWAT feed arteries (N; young=15, old=14). (G) Concentration response curves to insulin in BAT feed arteries in the absence or presence of the eNOS inhibitor, L-NAME (N; young=7, old=10). (H) Maximal insulin-stimulated vasodilation of BAT feed arteries (N; young=7, old=10). Data are shown as mean ± SEM. (C-H) *p<0.05 vs young with same treatment, †p<0.05 vs insulin within same group.

We next assessed the impact of aging on insulin-mediated vasodilation in AT feed arteries. Insulin-stimulated vasodilation was lower in all three AT feed arteries from old compared to young mice (all p<0.05) (Figure 1C, 1E, 1G). This reduction appears to result from a loss of NO bioavailability, as aging abolished the effect of L-NAME to reduce insulin-mediated dilation in arteries from all three AT depots (Figure 1C, 1E, 1G). Likewise, insulin-stimulated maximum dilation was lower in feed arteries from old compared to young mice in the pgWAT, scWAT and BAT (all <0.05) (Figure 1D, 1F, 1H). EC50 was higher in old pgWAT feed arteries and tended to be higher in scWAT and BAT feed arteries compared to young pgWAT, scWAT and BAT feed arteries, respectively (p<0.05, p=0.09, p=0.08) (Figure 1). Taken together, these data demonstrate tissue specificity in the dilatory response to insulin across AT depots and further that aging impairs dilation and NO bioavailability across all three depots.

Endothelium-independent dilation is not impacted by AT depot and/or aging in AT feed arteries.

To assess the endothelium-independent vasodilation, we measured the vasodilation in response to the inorganic NO donor, sodium nitroprusside (SNP). In response to SNP, AT feed arteries from all three depots dilated 85 to 95%, with no differences observed between depots (all p>0.05) (Figure 2). EC50 was not different between feed arteries from different AT depots (pgWAT: −7.96 ± 0.13; scWAT: −7.70 ± 0.11; BAT: −7.40 ± 0.11; all p>0.05). Aging did not alter SNP-mediated vasodilation in feed arteries from pgWAT, scWAT and BAT (p>0.05) (Figure 2). Likewise, there was no effect of age on sensitivity to SNP (EC50) in any of the AT feed arteries (all p>0.05) (Figure 2). Taken together, these results suggest that the impaired vasodilation observed in response to insulin with aging is mediated by endothelial dysfunction rather than smooth muscle dysfunction.

Figure 2: Sodium nitroprusside (SNP)-stimulated vasodilation in adipose tissue feed arteries from young and old mice.

Figure 2:

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(A) Concentration response curves to SNP in pgWAT feed arteries (N; young=11, old=18). (B) Maximal vasodilation to SNP in pgWAT feed arteries (N; young=11, old=18). (C) Concentration response curves to SNP in scWAT feed arteries (N; young=15, old=18). (D) Maximal vasodilation to SNP in scWAT feed arteries (N; young=15, old=18). (E) Concentration response curves to SNP in BAT feed arteries (N; young=8, old=16). (F) Maximal vasodilation to SNP in BAT feed arteries (N; young=8, old=16). Data are shown as mean ± SEM.

Advanced age modulates AT angiogenic capacity.

To assess the angiogenic capacity of different AT depots as well as the impact of aging, we performed an ex vivo sprouting assay and examined the number of tubes formed in a collagen matrix. In young mice, the scWAT vasculature demonstrated lower angiogenic sprout formation compared to pgWAT and BAT (both p<0.05), while the BAT vasculature demonstrated the highest angiogenic potential among three depots (p<0.05) (pgWAT: 1.88±0.20; scWAT 0.37±0.12; BAT: 2.96±0.61) (Figure 3). We next examined the impact of aging on angiogenic capacity and found that aging resulted in a reduction in angiogenic sprout formation in pgWAT and BAT (both p<0.05) (Figure 3A, 4C), but resulted in an increase in angiogenic sprout formation in the scWAT (p<0.05) (Figure 3B).

Figure 3: Angiogenic capacity of adipose tissues from young and old mice.

Figure 3:

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Number of angiogenic sprouts formed from (A) pgWAT, (B) scWAT and (C) BAT explants from young and old mice after 5 days of culture in a collagen matrix and treatment with vascular endothelial growth factor (VEGF: 0.5 mg/ml). (N; young=7-8, old=6-7). Data are shown as mean ± SEM. *p<0.05 vs young.

Discussion

In this study we aimed to determine; first, if endothelial function, assessed by both insulin-stimulated vasodilation and angiogenic capacity, differs across AT depots and; second, if aging differentially impacts endothelial function in these depots. This is the first study to examine insulin-stimulated vasodilation and angiogenic capacity across different AT depots and to elucidate potential age-related changes. The novel findings of this study are: (1) Insulin-stimulated vasodilation is lower in feed arteries from the pgWAT compared to the scWAT, but there is no difference between either the pgWAT and BAT or the scWAT and BAT, suggesting that circulating insulin may differentially impact vasodilation and potentially blood flow across the AT depots. (2) With aging, insulin-stimulated vasodilation is impaired in all three AT feed arteries and this impairment is endothelial-dependent and mediated through reduced NO bioavailability. (3) Among the three AT depots, angiogenic capacity is highest in the BAT, followed by the pgWAT and then the scWAT vasculature. (4) Advancing age resulted in a decrease in angiogenic capacity in pgWAT and BAT vasculature but in an increase in angiogenic capacity in scWAT vasculature. (5) The pgWAT demonstrated the highest tissue volume and mass followed by scWAT and BAT and advanced age led to a reduction in both the volume and mass of all three depots. Taken together, these findings suggest that insulin-stimulated vasodilation and angiogenic capacity differ across the vasculature of different AT depots. Advancing age impairs all of these functions except for angiogenesis in the scWAT, which is higher in old mice compared to young.

Skeletal muscle and ATs are the primary sites of insulin-stimulated glucose uptake and the vascular endothelium is a critical regulator in this process (DeFronzo & Tripathy, 2009; Satoh, 2014). Insulin has a multitude of roles in the vascular endothelium, contributing to the maintenance of metabolic homeostasis and, more specifically, contributing to glucose uptake. First, insulin leads to NO-mediated vasodilation, resulting in increases in local blood flow and thus provision of glucose to the site of clearance, such as skeletal muscle and ATs (Baron et al., 1995; Lambadiari, Triantafyllou, & Dimitriadis, 2015). Second, insulin signaling in the ECs facilitates its own trafficking through the EC barrier to the interstitium (Konishi et al., 2017; Kubota, Kubota, & Kadowaki, 2013). Insulin in the interstitial space leads to the phosphorylation of the canonical Akt signaling pathway leading to the translocation of glucose transporter Glut4 and glucose uptake (Petersen & Shulman, 2018). Numerous recent reports have demonstrated that genetic manipulation of insulin signaling in ECs impairs skeletal muscle glucose uptake leading to systemic metabolic dysfunction (Konishi et al., 2017; Kubota et al., 2011). While insulin-stimulated vasodilation in skeletal muscle is well established, the vasodilator responsiveness of AT arteries to insulin was not elucidated. In the present study, we found that feed arteries from all AT depots respond with approximately 40-50% maximum dilation to insulin and that this vasodilation was primarily mediated through NO. With advancing age, insulin-stimulated vasodilation and NO bioavailability was blunted in the feed arteries from all AT depots. In our previous study, we demonstrated blunted acetylcholine-mediated vasodilation and a reduced angiogenic capacity in the pgWAT that was associated with age-related metabolic dysfunction, especially glucose intolerance and insulin resistance, in this same mouse strain (Donato et al., 2014). Here, we extend these findings by demonstrating blunted insulin-stimulated vasodilation of AT feed arteries across all three major depots with aging, suggestive of endothelial insulin resistance. While this finding suggests that with aging there may be reduced blood flow to all three major AT depots in the postprandial state, the impact of this reduction in vasodilation on blood flow to, and function of, these AT depots require further elucidation.

Angiogenesis is a process of neovessel formation that contributes to AT vascular density, perfusion, and consequently oxygen and nutrient supply (Rupnick et al., 2002). Declines in angiogenic capacity may lead to inadequate AT vascular density, resulting in AT hypoxia, inflammation and metabolic dysfunction (Gregor & Hotamisligil, 2007; Yilmaz & Hotamisligil, 2013; Zhang et al., 2011). VEGF signaling plays a critical role in neovessel formation and contributes to the maintenance of appropriate vascularity (Shibuya, 2011). In the present study, we found that in healthy young mice, the BAT vasculature has the highest VEGF-stimulated angiogenic potential followed by pgWAT and scWAT. BAT and pgWAT from old mice demonstrated lower VEGF-stimulated angiogenic potential, while the scWAT demonstrated higher angiogenic potential compared to young controls. These results suggest that reduced VEGF signaling may contribute to an age-related decline in AT vascularity of the pgWAT and the BAT, which may contribute to AT inflammation and metabolic dysfunction that has been well described in aging (Donato et al., 2014; Palmer & Kirkland, 2016). The implications of an increased angiogenic capacity in the scWAT with aging are unclear but may be a compensatory adaptation. In human aging, there is a redistribution of lipids from the subcutaneous to the visceral stores from middle to older age (Palmer & Kirkland, 2016), and if the observed increased angiogenic capacity also occurs with human aging, it may serve to limit this loss of lipid storage in the subcutaneous depots. Likewise, as aging is associated with systemic metabolic dysfunction, an upregulation of angiogenesis in the scWAT may be a mechanism by which the organism is attempting to increase nutrient supply in the face of diminished vasodilator responsiveness. Such an adaptation may be beneficial systemically by helping to reduce circulating lipids and by increasing the volume of this more protective AT depot. Whether the scWAT maintains its “protective” phenotype under these conditions, however, requires further study. Another possible explanation for the differential findings between the AT depots may stem from the growth factor used in this study. Here we examined in vitro angiogenic capacity in response to VEGF, but in a physiological setting angiogenesis is more complex and stimulated by a number of growth factors. It may be that there is differential sensitivity of the individual AT depots to individual growth factors and/or that these may be impacted differentially by aging. Therefore, future studies are warranted to examine the vascular density, blood flow and in vivo angiogenesis in response to a variety of growth factors both across AT depots and in the setting of aging.

Aging results in a decline in AT function that is associated with hypoxia, inflammation, metabolic dysfunction and declines in tissue mass (Palmer & Kirkland, 2016). Adequate blood flow and thus continuous provision of oxygen and nutrients are the prerequisites for organ function (Gregor & Hotamisligil, 2007; Yilmaz & Hotamisligil, 2013; Zhang et al., 2011). Insulin action on vascular endothelium is critical for vasodilation and thus blood flow leading to the supply of nutrients and oxygen and trafficking of insulin through EC barrier leading to glucose and fatty acid uptake (Baron et al., 1995; Kubota et al., 2013). In addition to vasodilation, angiogenesis plays a critical role in maintaining AT vascularity and blood flow (Sun, Kusminski, & Scherer, 2011). An impairment in these processes may lead to inadequate nutrient supply resulting in impaired metabolic function and reduced tissue mass. The present study demonstrates that aging impairs both insulin-stimulated endothelium-dependent dilation and angiogenesis in AT arteries. Therefore, results from this study support the conclusion that age-related impairments of endothelial function in the AT are potential contributing factors to hypoxia, inflammation and metabolic dysfunction and declines in tissue mass.

Limitations and Future Directions

This study has some limitations that may be addressed in future studies. For example, this study cannot conclude if age-related impairments in insulin-stimulated vasodilation and angiogenic capacity are the cause or consequence of AT dysfunction. Genetic manipulation of the vascular endothelium related to its vasodilatory and angiogenic functions could be performed to determine if blunting insulin stimulated vasodilation and impaired angiogenesis in AT arteries phenocopies AT aging. Although aging is well known to be associated with AT hypoxia, metabolic dysfunction and inflammation, these were not evaluated in the present study. Future studies should elucidate the direct role of manipulating endothelial function on these well-established characteristics of aged AT. Furthermore, in this study, we found that the impact of aging on angiogenesis is similar between pgWAT and BAT vasculature but opposite in the scWAT vasculature, the implications of this differential effect of aging on angiogenesis among AT depots requires further elucidation.

Grant Support:

This work was supported by National Institutes of Health awards: R01 AG060395, R01 AG048366, R00 AT010017, and Veteran's Affairs Merit Review Awards I01 BX002151 and I01 BX004492 from the United States (U.S.) Department of Veterans Affairs Biomedical Laboratory Research and Development Service. The contents do not represent the views of the U.S. Department of Veterans Affairs, the National Institutes of Health or the United States Government.

Footnotes

Disclosures: None.

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