Most studies in adults suggest that acute glucose consumption induces a transient impairment in endothelial function. We hypothesized that obese youth would demonstrate reduced endothelial function and increased inflammation and oxidative stress following acute glucose ingestion and that transient elevations in plasma glucose would correlate with endothelial dysfunction, inflammation, and oxidative stress. Thirty-four obese (BMI ≥95th percentile) children and adolescents (age 12.4 ± 2.6 years; BMI = 37.9 ± 6.7 kg/m2; 50% females) underwent measurement of endothelial function (reactive hyperemic index (RHI)), glucose, insulin, C-reactive protein (CRP), interleukin-6 (IL-6), circulating oxidized low-density lipoprotein (oxLDL), and myeloperoxidase (MPO) in a fasting state and at 1- and 2-h following glucose ingestion. Repeated measures ANOVA with Tukey post-tests and Pearson correlations were performed. Glucose and insulin levels significantly increased at 1- and 2-h (all P values < 0.001). Compared to baseline, there were no statistically significant differences in 1- and 2-h RHI, CRP, IL-6, and oxLDL. However, MPO significantly decreased at the 1- (P < 0.05) and 2-h (P < 0.001) time points. At the 1-h time point, glucose level was significantly inversely correlated with RHI (r = −0.40, P < 0.05) and at the 2-h time point, glucose level was positively correlated with MPO (r = 0.40, P < 0.05). An acute oral glucose load does not reduce endothelial function or increase levels of inflammation or oxidative stress in obese youth. However, associations of postprandial hyperglycemia with endothelial function and oxidative stress may have implications for individuals with impaired glucose tolerance or frank type 2 diabetes.
Endothelial dysfunction is one of the earliest detectable signs of the initiation of cardiovascular disease and predicts subsequent atherosclerosis (1) and future cardiovascular events (2,3,4,5). Healthy vascular endothelial cells exert tight control over arterial tone by producing and regulating vasodilating factors such as nitric oxide, a free radical that stimulates vasorelaxation of the underlying smooth muscle cells (6). In addition to stimulating vasodilation, nitric oxide performs a myriad of antiatherosclerotic functions such as interfering with monocyte adhesion to the arterial wall (7), inhibiting smooth muscle cell proliferation (8), and decreasing platelet aggregation (9). The endothelium is fundamentally involved with the atherosclerotic process in part due to its location. Endothelial cells are strategically located at the interface of the lumen and the artery wall, putting them in direct contact with substances carried in the blood that may interfere with their proper functioning.
Glucose is one such factor that can directly affect the function of the endothelium. Most, but not all, studies in adults suggest that transient (up to 3 h) elevations in blood glucose acutely impair endothelial function. Studies in adults with either impaired glucose tolerance or type 2 diabetes mellitus have consistently reported reductions in postprandial endothelial function (10,11,12,13). However, results have been mixed regarding whether healthy adults experience transient endothelial dysfunction in the postprandial setting. Some studies report no change in endothelial function with acute hyperglycemia (14,15,16,17), while others report impairment (11,18,19,20,21). The mechanisms by which acute hyperglycemia instigates endothelial dysfunction is not completely clear but is likely related to elevations in inflammation and oxidative stress (10,11,19,22,23,24), which are known to interfere with nitric oxide production and bioavailability.
The effect of postprandial glucose excursions on endothelial function has not been well described in children and adolescents. To our knowledge, a previous study from our research group is the only one to have addressed this question. We reported that conduit artery endothelial function, as measured by brachial artery flow-mediated dilation, was not impaired following an oral glucose challenge in overweight (BMI ≥85th percentile) or normal-weight children and adolescents (25). However, postprandial glucose excursions and endothelial dysfunction are generally more pronounced at higher levels of adiposity (26,27). Therefore, we conducted the present study to evaluate postprandial endothelial function, inflammation and oxidative stress in obese (BMI ≥95th percentile) children and adolescents. We hypothesized that obese youth would demonstrate acute endothelial dysfunction and elevations in inflammation and oxidative stress following glucose ingestion and that transient elevations in blood glucose would correlate with endothelial dysfunction and plasma biomarkers of inflammation and oxidative stress.
Methods and Procedures
Thirty-four obese (BMI ≥95th percentile based on gender and age) children and adolescents were consecutively enrolled from a pediatric weight management clinic (no exclusion criteria were used). Participants had newly entered the program and had not yet initiated any behavioral or drug therapies. Measures were obtained after participants had been fasting for at least 10 h. The protocol was approved by the University of Minnesota institutional review board and consent/assent was obtained from parents/participants.
Anthropometric measures and blood pressure
Height and weight were obtained using a standard stadiometer and electronic scale, respectively. BMI was calculated as weight (kg)/height (m2). Seated blood pressure was obtained after 5 min of quiet rest, on the right arm using an automatic sphygmomanometer.
Oral glucose tolerance test and blood analyses
A standard oral glucose tolerance test, consisting of a 75-g glucose load, was performed after fasting and blood was collected for measurement of lipids, glucose, insulin, C-reactive protein (CRP), interleukin-6 (IL-6), circulating oxidized low-density lipoprotein (oxLDL), and myeloperoxidase (MPO). Blood was collected every 30 min for a period of 2 h for measurement of glucose and insulin and at the 1- and 2-h time points for measurement of CRP, IL-6, oxLDL, and MPO. Fasting lipid profile, glucose, and insulin assays were conducted with standard procedures at the Fairview Diagnostic Laboratories, Fairview-University Medical Center (Minneapolis, MN), a Center for Disease Control and Prevention-certified laboratory. Insulin resistance was estimated by Matsuda index (28) and homeostasis model assessment for insulin resistance (29). Blood samples for plasma biomarkers were centrifuged and plasma was stored at −80 °C for a batched analysis at the end of the study. CRP, IL-6, oxLDL, and MPO levels were measured by enzyme-linked immunosorbent assay in the University of Minnesota Cytokine Reference Laboratory (Minneapolis, MN) (Clinical Laboratory Improvement Amendments licensed).
Measurement of endothelial function
Endothelial function was measured noninvasively by digital reactive hyperemia (EndoPAT 2000; Itamar Medical, Caesarea, Israel) at baseline (prior to glucose ingestion), 1- and 2-h following the glucose load. Endothelial function as assessed by this technique is nitric oxide-dependent (30), associated with coronary artery blood flow (31) and multiple cardiovascular risk factors (32), and independently predicts future cardiovascular events (5). After 10 min of quiet rest in the supine position, finger probes were placed on the index fingers of both hands to measure baseline and reactive hyperemic pulse amplitude. The probes inflate to apply a uniform pressure (10 mm Hg less than diastolic blood pressure) on the fingers and detect small pulse volume changes throughout the cardiac cycle. Following the collection of 5 min of baseline data, a blood pressure cuff on the upper forearm (just below the elbow) was inflated to a suprasystolic level for 5 min. Following cuff release, the change in pulse amplitude during reactive hyperemia was measured for 5 min. The ratio of the hyperemic and the baseline pulse amplitude (corrected for the same ratio on the control finger) was calculated and expressed as the reactive hyperemic index (RHI). Reproducibility of this technique in children is excellent. The mean difference in RHI measured 1 week apart was 0.12 and within subject variation was 0.16 (33).
Repeated measures ANOVA, with Tukey post-tests, were used to compare RHI, CRP, IL-6, oxLDL, and MPO at baseline, 1-h, and 2-h during the oral glucose tolerance test. Pearson correlation analyses were performed to assess associations of glucose with RHI, CRP, IL-6, oxLDL, and MPO at 1-h and 2-h during the oral glucose tolerance test. Data are presented as mean ± s.d. P values <0.05 were considered statistically significant.
Clinical characteristics of participants are presented in Table 1. Mean age of the participants was 12.4 ± 2.6 years (50% females) and mean BMI was 37.9 ± 6.7 kg/m2. Known clinical diagnoses included 21 (62%) participants with acanthosis nigricans, five (15%) with hypertension, five (15%) with dyslipidemia, and three (9%) with sleep apnea. In addition, five (15%) of the participants had impaired glucose tolerance and none had impaired fasting glucose or type 2 diabetes mellitus. Compared to baseline (79.9 ± 6.4 mg/dl), glucose significantly increased at the 1-h (124.0 ± 30.4 mg/dl, P < 0.001) and 2-h (114.2 ± 21.0 mg/dl, P < 0.001) time points (Figure 1a). Compared to baseline (17.4 ± 11.5 mU/l), insulin significantly increased at the 1-h (107.5 ± 73.0 mU/l, P < 0.0001) and 2-h (108.4 ± 75.3 mU/l, P < 0.0001) time points (Figure 1b). Compared to baseline (1.73 ± 0.5), there were no significant differences in the 1-h (1.59 ± 0.5) and 2-h (1.64 ± 0.5) RHI (Figure 2). Similarly, there were no differences between baseline and 1-h and 2-h CRP (5.8 ± 5.1 mg/l vs. 5.4 ± 5.1 mg/l vs. 5.1 ± 4.4 mg/l, respectively), IL-6 (2.0 ± 0.9 pg/ml vs. 1.9 ± 1.0 pg/ml vs. 1.8 ± 0.8 pg/ml, respectively), and oxLDL (66.3 ± 13.1 U/l vs. 62.1 ± 11.3 U/l vs. 61.0 ± 11.3 U/l, respectively). Compared to baseline (67.4 ± 22.0 µg/l), MPO significantly decreased at the 1-h (60.8 ± 18.5 µg/l, P < 0.05) and 2-h (54.7 ± 20.3 µg/l, P < 0.001) time points. At the 1-h time point, glucose level was significantly inversely correlated with RHI (r = −0.40, P < 0.05). At the 2-h time point, glucose level was significantly correlated with MPO (r = 0.40, P < 0.05).
Table 1. Clinical characteristics
The main finding of this study is that obese children and adolescents do not demonstrate acute endothelial dysfunction or elevations in inflammation and oxidative stress following an oral glucose challenge. However, significant associations exist between the degree of postprandial hyperglycemia and both endothelial function and oxidative stress. The latter findings suggest that children with more severe blood glucose excursions in the postprandial setting, such as those observed in individuals with impaired glucose tolerance or frank type 2 diabetes, may experience a transient period (up to 2 h) of endothelial dysfunction and oxidative stress following carbohydrate consumption.
The current findings are consistent with our previous study in overweight children and adolescents (25), which showed no reduction in conduit artery endothelial function, as measured by brachial artery flow-mediated dilation, following an acute glucose challenge. However, the current study expands our knowledge in this area in a few important ways. First, participants in this study had much higher levels of adiposity than in the previous study (mean BMI = 37.9 vs. 29.3 kg/m2, respectively), thereby extending our earlier findings to an obese pediatric population. Indeed, most of the participants in the current study were classified as extremely obese (BMI ≥1.2 times the 95th percentile) (34,35). Second, we evaluated endothelial function in the microvasculature, a vascular bed not explored in our previous study, and have shown that resistance artery endothelial function is similarly unaffected by acute hyperglycemia. Finally, we measured multiple plasma biomarkers of inflammation and oxidative stress in an attempt to address potential mechanisms of hyperglycemia-mediated endothelial dysfunction. Contrary to most studies that have addressed postprandial endothelial activation in adults (10,11,12,13,18,19,20,21,22,23,24), our data, showing that endothelial function remained intact and levels of inflammation and oxidative stress did not increase following a glucose challenge, provide evidence that this phenomenon does not occur in obese children and adolescents and suggests that it may not develop until later in life.
Despite the lack of a significant change in postprandial endothelial function and levels of inflammation and oxidative stress, we observed a significant inverse association between blood glucose level and RHI at the 1-h time point and a significant positive association between blood glucose level and MPO at the 2-h time point. The relationship between the magnitude of change in postprandial glucose level and both endothelial dysfunction and oxidative stress may have implications for youth with impaired glucose tolerance and/or type 2 diabetes mellitus. By definition, these individuals have an exaggerated glycemic response to oral glucose ingestion and therefore may experience significant reductions in endothelial function and increases in levels of oxidative stress. Although yet to be explored in children, data in adults with type 2 diabetes mellitus suggest that acute glucose fluctuations instigate higher levels of oxidative stress than chronic sustained hyperglycemia (36).
Contrary to our hypothesis, we found that levels of MPO significantly decreased during the glucose challenge. However, a similar observation was reported in a recent study of women with and without type 2 diabetes mellitus (37). In this study, MPO levels significantly decreased following two separate meal challenges, one consisting of a high percentage of carbohydrates and the other consisting of a high percentage of fat. Since it is unclear why MPO levels seem to decrease in the postprandial setting, this may be an important topic of further investigation.
This study had several limitations that should be noted. Tanner stage was not assessed on all subjects. Therefore, we were unable to compare the effects of pubertal status on postprandial endothelial function, inflammation, and oxidative stress. The generalizability of our results may be limited since subjects in our study were recruited from a pediatric weight management clinic. Factors other than obesity (such as family history, dyslipidemia, hypertension, etc.) may explain why these patients were referred for specialty care; therefore, subjects in our study may not be representative of the general obese pediatric population. Additional characteristics, such as family history of cardiovascular disease and type 2 diabetes and dietary/physical activity patterns, were not collected. In addition, only a limited number of inflammation and oxidative stress markers were measured.
In conclusion, findings from this study suggest that oral consumption of glucose does not acutely impair endothelial function or increase levels of inflammation or oxidative stress in obese children and adolescents. The results provide evidence that arteries may retain the ability to properly regulate blood flow and dilatory capacity within the postprandial setting during childhood, even in the context of obesity, and that a window of opportunity may exist during this time to institute interventions aimed at maintaining this status. However, the degree of postprandial hyperglycemia is associated with endothelial function and oxidative stress, which may have implications for youth with impaired glucose tolerance and/or type 2 diabetes mellitus. Further work will be required to explore whether children and adolescents with these conditions experience postprandial endothelial dysfunction and will be particularly relevant given the increasing prevalence of these conditions in youth.
The authors declared no conflict of interest. Funding for this study was provided by the University of Minnesota Vikings Children's Fund (A.S.K.) and Minnesota Medical Foundation (A.S.K.). Mercodia, generously provided discounted enzyme-linked immunosorbent assay kits for the oxidized low-density lipoprotein and myeloperoxidase analyses.