Joyce M. Richey

Angiotensin II induces insulin resistance independent of changes in interstitial insulin

We set out to examine whether angiotensin-driven hypertension can alter insulin action and whether these changes are reflected as changes in interstitial insulin (the signal to which insulin-sensitive cells respond to increase glucose uptake). To this end, we measured hemodynamic parameters, glucose turnover, and insulin dynamics in both plasma and interstitial fluid (lymph) during hyperinsulinemic euglycemic clamps in anesthetized dogs, with or without simultaneous infusions of angiotensin II (ANG II). Hyperinsulinemia per se failed to alter mean arterial pressure, heart rate, or femoral blood flow. ANG II infusion resulted in increased mean arterial pressure (68 ± 16 to 94 ± 14 mmHg, P < 0.001) with a compensatory decrease in heart rate (110 ± 7 vs. 86 ± 4 mmHg, P < 0.05). Peripheral resistance was significantly increased by ANG II from 0.434 to 0.507 mmHg ⋅ ml-1 ⋅ min ( P < 0.05). ANG II infusion increased femoral artery blood flow (176 ± 4 to 187 ± 5 ml/min, P < 0.05) and resulted in additional increases in both plasma and lymph insulin (93 ± 20 to 122 ± 13 μU/ml and 30 ± 4 to 45 ± 8 μU/ml, P < 0.05). However, glucose uptake was not significantly altered and actually had a tendency to be lower (5.9 ± 1.2 vs. 5.4 ± 0.7 mg ⋅ kg-1 ⋅ min-1, P > 0.10). Mimicking of the ANG II-induced hyperinsulinemia resulted in an additional increase in glucose uptake. These data imply that ANG II induces insulin resistance by an effect independent of a reduction in interstitial insulin.We set out to examine whether angiotensin-driven hypertension can alter insulin action and whether these changes are reflected as changes in interstitial insulin (the signal to which insulin-sensitive cells respond to increase glucose uptake). To this end, we measured hemodynamic parameters, glucose turnover, and insulin dynamics in both plasma and interstitial fluid (lymph) during hyperinsulinemic euglycemic clamps in anesthetized dogs, with or without simultaneous infusions of angiotensin II (ANG II). Hyperinsulinemia per se failed to alter mean arterial pressure, heart rate, or femoral blood flow. ANG II infusion resulted in increased mean arterial pressure (68 +/- 16 to 94 +/- 14 mmHg, P < 0. 001) with a compensatory decrease in heart rate (110 +/- 7 vs. 86 +/- 4 mmHg, P < 0.05). Peripheral resistance was significantly increased by ANG II from 0.434 to 0.507 mmHg. ml(-1). min (P < 0.05). ANG II infusion increased femoral artery blood flow (176 +/- 4 to 187 +/- 5 ml/min, P < 0.05) and resulted in additional increases in both plasma and lymph insulin (93 +/- 20 to 122 +/- 13 microU/ml and 30 +/- 4 to 45 +/- 8 microU/ml, P < 0.05). However, glucose uptake was not significantly altered and actually had a tendency to be lower (5.9 +/- 1.2 vs. 5.4 +/- 0.7 mg. kg(-1). min(-1), P > 0.10). Mimicking of the ANG II-induced hyperinsulinemia resulted in an additional increase in glucose uptake. These data imply that ANG II induces insulin resistance by an effect independent of a reduction in interstitial insulin.

Direct Administration of Insulin Into Skeletal Muscle Reveals That the Transport of Insulin Across the Capillary Endothelium Limits the Time Course of Insulin to Activate Glucose Disposal

OBJECTIVE—Intravenous insulin infusion rapidly increases plasma insulin, yet glucose disposal occurs at a much slower rate. This delay in insulins action may be related to the protracted time for insulin to traverse the capillary endothelium. An increased delay may be associated with the development of insulin resistance. The purpose of the present study was to investigate whether bypassing the transendothelial insulin transport step and injecting insulin directly into the interstitial space would moderate the delay in glucose uptake observed with intravenous administration of the hormone. RESEARCH DESIGN AND METHODS—Intramuscular injections of saline (n = 3) or insulin (n = 10) were administered directly into the vastus medialis of anesthetized dogs. Injections of 0.3, 0.5, 0.7, 1.0, and 3.0 units insulin were administered hourly during a basal insulin euglycemic glucose clamp (0.2mU · min−1 · kg−1). RESULTS—Unlike the saline group, each incremental insulin injection caused interstitial (lymph) insulin to rise within 10 min, indicating rapid diffusion of the hormone within the interstitial matrix. Delay in insulin action was virtually eliminated, indicated by immediate dose-dependent increments in hindlimb glucose uptake. Additionally, bypassing insulin transport by direct injection into muscle revealed a fourfold greater sensitivity to insulin of in vivo muscle tissue than previously reported from intravenous insulin administration. CONCLUSIONS—Our results indicate that the transport of insulin to skeletal muscle is a rate-limiting step for insulin to activate glucose disposal. Based on these results, we speculate that defects in insulin transport across the endothelial layer of skeletal muscle will contribute to insulin resistance.

Reduced access to insulin-sensitive tissues in dogs with obesity secondary to increased fat intake

Physiological hyperinsulinemia provokes hemodynamic actions and augments access of macromolecules to insulin-sensitive tissues. We investigated whether induction of insulin resistance by a hypercaloric high-fat diet has an effect on the extracellular distribution of macromolecules to insulin-sensitive tissues. Male mongrel dogs were randomly selected into two groups: seven dogs were fed an isocaloric control diet (∼3,900 kcal, 35% from fat), and six dogs were fed a hypercaloric high-fat diet (∼5,300 kcal, 54% from fat) for a period of 12 weeks. During hyperinsulinemic-euglycemic clamps, we determined transport parameters and distribution volumes of [14C]inulin by applying a three-compartment model to the plasma clearance data of intravenously injected [14C]inulin (0.8 μCi/kg). In another study with direct cannulation of the hindlimb skeletal muscle lymphatics, we investigated the effect of physiological hyperinsulinemia on the appearance of intravenously injected [14C]inulin in skeletal muscle interstitial fluid and compared the effect of insulin between control and high-fat diet groups. The hypercaloric high-fat diet resulted in significant weight gain (18%; P < 0.001) associated with marked increases of subcutaneous (140%; P < 0.001) and omental (83%; P < 0.001) fat depots, as well as peripheral insulin resistance, measured as a significant reduction of insulin-stimulated glucose uptake during clamps (−35%; P < 0.05). Concomitantly, we observed a significant reduction of the peripheral distribution volume of [14C]inulin (−26%; P < 0.05), whereas the vascular distribution volume and transport and clearance parameters did not change as a cause of the diet. The second study directly confirmed our findings, suggesting a marked reduction of insulin action to stimulate access of macromolecules to insulin-sensitive tissues (control diet 32%, P < 0.01; high-fat diet 18%, NS). The present results indicate that access of macromolecules to insulin-sensitive tissues is impaired during diet-induced insulin resistance and suggest that the ability of insulin itself to stimulate tissue access is diminished. We speculate that the observed diet-induced defects in stimulation of tissue perfusion contribute to the development of peripheral insulin resistance.

Large Size Cells in the Visceral Adipose Depot Predict Insulin Resistance in the Canine Model

Adipocyte size plays a key role in the development of insulin resistance. We examined longitudinal changes in adipocyte size and distribution in visceral (VIS) and subcutaneous (SQ) fat during obesity‐induced insulin resistance and after treatment with CB‐1 receptor antagonist, rimonabant (RIM) in canines. We also examined whether adipocyte size and/or distribution is predictive of insulin resistance. Adipocyte morphology was assessed by direct microscopy and analysis of digital images in previously studied animals 6 weeks after high‐fat diet (HFD) and 16 weeks of HFD + placebo (PL; n = 8) or HFD + RIM (1.25 mg/kg/day; n = 11). At 6 weeks, mean adipocyte diameter increased in both depots with a bimodal pattern only in VIS. Sixteen weeks of HFD+PL resulted in four normally distributed cell populations in VIS and a bimodal pattern in SQ. Multilevel mixed‐effects linear regression with random‐effects model of repeated measures showed that size combined with share of adipocytes >75 µm in VIS only was related to hepatic insulin resistance. VIS adipocytes >75 µm were predictive of whole body and hepatic insulin resistance. In contrast, there was no predictive power of SQ adipocytes >75 µm regarding insulin resistance. RIM prevented the formation of large cells, normalizing to pre‐fat status in both depots. The appearance of hypertrophic adipocytes in VIS is a critical predictor of insulin resistance, supporting the deleterious effects of increased VIS adiposity in the pathogenesis of insulin resistance.

Fructose perfusion in rat mesenteric arteries impairs endothelium-dependent vasodilation

We demonstrated that the fructose-induced hypertensive rat, representative of the principal metabolic abnormalities found in a majority of hypertensive patients, i.e. hypertriglyceridemia, hyperinsulinemia and insulin resistance (Syndrome X), is associated with an impaired response to endothelium-dependent vasodilators and that fructose may directly contribute to this impairment. Twelve male Wistar rats were divided into two groups, one given 10% fructose (n=6); the other no fructose (n=6) for 40 days in the drinking water. Systolic blood pressure was measured via the tail cuff method. Perfusion pressure responses to acetylcholine, were measured in the isolated perfused mesenteric vascular bed. Constrictor or dilator responses were measured as increases or decreases, respectively, of the perfusion pressure at a constant flow (4 ml/min). Fructose-fed rats had significantly higher blood pressure, insulin and triglyceride levels than control animals. In phenylephrine constricted beds, the endothelium-dependent dilatation to acetylcholine (0.001 to 1 micromol) was attenuated in the fructose-fed group compared to control animals. Whether this abnormality results from the syndromes (hyperinsulinemia, hypertension and hypertriglyceridemia) associated with the fructose-fed animal model is unknown. We therefore hypothesized that fructose can impair the endothelium-dependent vasodilator response. This was evaluated by perfusing mesenteric arteries from normal rats with control mannitol (40 mM) or fructose (40 mM). Endothelium-dependent dilation to acetylcholine was impaired in fructose-perfused mesenteric arteries. Indomethacin restored the vasodilator response to acetylcholine, suggesting that a cyclooxygenase derivative mediates the impaired response. Thus, we conclude that fructose can contribute to the impaired endothelium-dependent response in the fructose-induced hypertensive rat model.

Rimonabant prevents additional accumulation of visceral and subcutaneous fat during high-fat feeding in dogs

We investigated whether rimonabant, a type 1 cannabinoid receptor antagonist, reduces visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) in dogs maintained on a hypercaloric high-fat diet (HHFD). To determine whether energy expenditure contributed to body weight changes, we also calculated resting metabolic rate. Twenty male dogs received either rimonabant (1.25 mg.kg(-1).day(-1), orally; n = 11) or placebo (n = 9) for 16 wk, concomitant with a HHFD. VAT, SAT, and nonfat tissue were measured by magnetic resonance imaging. Resting metabolic rate was assessed by indirect calorimetry. By week 16 of treatment, rimonabant dogs lost 2.5% of their body weight (P = 0.029), whereas in placebo dogs body weight increased by 6.2% (P < 0.001). Rimonabant reduced food intake (P = 0.027), concomitant with a reduction of SAT by 19.5% (P < 0.001). In contrast with the VAT increase with placebo (P < 0.01), VAT did not change with rimonabant. Nonfat tissue remained unchanged in both groups. Body weight loss was not associated with either resting metabolic rate (r(2) = 0.24; P = 0.154) or food intake (r(2) = 0.24; P = 0.166). In conclusion, rimonabant reduced body weight together with a reduction in abdominal fat, mainly because of SAT loss. Body weight changes were not associated with either resting metabolic rate or food intake. The findings provide evidence of a peripheral effect of rimonabant to reduce adiposity and body weight, possibly through a direct effect on adipose tissue.

Consistency of the Disposition Index in the Face of Diet Induced Insulin Resistance: Potential Role of FFA

Objective Insulin resistance induces hyperinsulinemic compensation, which in turn maintains almost a constant disposition index. However, the signal that gives rise to the hyperinsulinemic compensation for insulin resistance remains unknown. Methods In a dog model of obesity we examined the possibility that potential early-week changes in plasma FFA, glucose, or both could be part of a cascade of signals that lead to compensatory hyperinsulinemia induced by insulin resistance. Results Hypercaloric high fat feeding in dogs resulted in modest weight gain, and an increase in adipose tissue with no change in the non-adipose tissue size. To compensate for the drop in insulin sensitivity, there was a significant rise in plasma insulin, which can be attributed in part to a decrease in the metabolic clearance rate of insulin and increased insulin secretion. In this study we observed complete compensation for high fat diet induced insulin resistance as measured by the disposition index. The compensatory hyperinsulinemia was coupled with significant changes in plasma FFAs and no change in plasma glucose. Conclusions We postulate that early in the development of diet induced insulin resistance, a change in plasma FFAs may directly, through signaling at the level of β-cell, or indirectly, by decreasing hepatic insulin clearance, result in the observed hyperinsulinemic compensation.

Hepatic insulin clearance is the primary determinant of insulin sensitivity in the normal dog.

Insulin resistance is a powerful risk factor for Type 2 diabetes and a constellation of chronic diseases, and is most commonly associated with obesity. We examined if factors other than obesity are more substantial predictors of insulin sensitivity under baseline, nonstimulated conditions.

CRP Is Related to Higher Leptin Levels in Minority Peripubertal Females Regardless of Adiposity Levels

Overweight is related to higher levels of C‐reactive protein (CRP) and leptin, which have been independently associated with increased risk for diabetes, cardiovascular disease, and the metabolic syndrome. Elevated CRP may trigger leptin resistance by inhibiting the binding of leptin to its receptors. We cross‐sectionally examined the relationship between CRP, leptin, BMI z‐score, percent body fat (%BF) assessed by air plethysmography (BodPod), and insulin sensitivity (SI) and acute insulin response (AIRg) measured by intravenous glucose tolerance test in 51 Latina and African‐American females (77% Latina), mean age 9.2 (±0.9) years, at either Tanner Pubertal Stage (TPS) 1 (n = 25) or TPS 2 (n = 26). Females at TPS 2 had higher BMI z‐scores, %BF (23% ± 10.1 vs. 30% ± 10.0, P = 0.02), AIRg (976.7 ± 735.2 vs. 1555.3 ± 1,223 µIU/ml, P = 0.05), fasting insulin (11.0 ± 10.8 vs. 17.2 ± 13.6 µlU/ml, P = 0.00) and leptin levels (11.0 ± 7.1 vs. 19.6 ± 10.9 ng/ml, P < 0.001) than those at TPS 1. There were no ethnic differences in any of the measured variables. CRP was positively correlated with BMI z‐score (P = 0.001), %BF (P = 0.006), fasting insulin and AIRg (P = 0.02), and fasting leptin (P = 0.00), and negatively correlated with SI (P = 0.05). A linear regression model showed that CRP independently explained 10% (P = 0.00) of the variance in leptin after adjusting %BF, TPS, ethnicity, habitual physical activity and SI. Hence, low‐grade inflammation may contribute to prolonged leptin exposure and leptin resistance, even in healthy children.

Failure of Homeostatic Model Assessment of Insulin Resistance to Detect Marked Diet-Induced Insulin Resistance in Dogs

Accurate quantification of insulin resistance is essential for determining efficacy of treatments to reduce diabetes risk. Gold-standard methods to assess resistance are available (e.g., hyperinsulinemic clamp or minimal model), but surrogate indices based solely on fasting values have attractive simplicity. One such surrogate, the homeostatic model assessment of insulin resistance (HOMA-IR), is widely applied despite known inaccuracies in characterizing resistance across groups. Of greater significance is whether HOMA-IR can detect changes in insulin sensitivity induced by an intervention. We tested the ability of HOMA-IR to detect high-fat diet–induced insulin resistance in 36 healthy canines using clamp and minimal model analysis of the intravenous glucose tolerance test (IVGTT) to document progression of resistance. The influence of pancreatic function on HOMA-IR accuracy was assessed using the acute insulin response during the IVGTT (AIRG). Diet-induced resistance was confirmed by both clamp and minimal model (P < 0.0001), and measures were correlated with each other (P = 0.001). In striking contrast, HOMA-IR ([fasting insulin (μU/mL) × fasting glucose (mmol)]/22.5) did not detect reduced sensitivity induced by fat feeding (P = 0.22). In fact, 13 of 36 animals showed an artifactual decrease in HOMA-IR (i.e., increased sensitivity). The ability of HOMA-IR to detect diet-induced resistance was particularly limited under conditions when insulin secretory function (AIRG) is less than robust. In conclusion, HOMA-IR is of limited utility for detecting diet-induced deterioration of insulin sensitivity quantified by glucose clamp or minimal model. Caution should be exercised when using HOMA-IR to detect insulin resistance when pancreatic function is compromised. It is necessary to use other accurate indices to detect longitudinal changes in insulin resistance with any confidence.