Renal and Vascular Effects of Combined SGLT2 and Angiotensin-Converting Enzyme Inhibition.

BACKGROUND: The cardiorenal effects of sodium-glucose cotransporter 2 inhibition (empagliflozin 25 mg QD) combined with angiotensin-converting enzyme inhibition (ramipril 10 mg QD) were assessed in this mechanistic study in patients with type 1 diabetes with potential renal hyperfiltration. METHODS: Thirty patients (out of 31 randomized) completed this double-blind, placebo-controlled, crossover trial. Recruitment was stopped early because of an unexpectedly low proportion of patients with hyperfiltration. Measurements were obtained after each of the 6 treatment phases over 19 weeks: (1) baseline without treatment, (2) 4-week run-in with ramipril treatment alone, (3) 4-week combined empagliflozin-ramipril treatment, (4) a 4-week washout, (5) 4-week combined placebo-ramipril treatment, and (6) 1-week follow-up. The primary end point was glomerular filtration rate (GFR) after combination treatment with empagliflozin-ramipril compared with placebo-ramipril. GFR was corrected for ramipril treatment alone before randomization. At the end of study phase, the following outcomes were measured under clamped euglycemia (4 to 6 mmol/L): inulin (GFR) and para-aminohippurate (effective renal plasma flow) clearances, tubular sodium handling, ambulatory blood pressure, arterial stiffness, heart rate variability, noninvasive cardiac output monitoring, plasma and urine biochemistry, markers of the renin-angiotensin-aldosterone system, and oxidative stress. RESULTS: Combination treatment with empagliflozin-ramipril resulted in an 8 mL/min/1.73 m2 lower GFR compared with placebo-ramipril treatment (P=0.0061) without significant changes to effective renal plasma flow. GFR decrease was accompanied by a 21.3 mL/min lower absolute proximal fluid reabsorption rate (P=0.0092), a 3.1 mmol/min lower absolute proximal sodium reabsorption rate (P=0.0056), and a 194 ng/mmol creatinine lower urinary 8-isoprostane level (P=0.0084) relative to placebo-ramipril combination treatment. Sodium-glucose cotransporter 2 inhibitor/angiotensin-converting enzyme inhibitor combination treatment resulted in additive blood pressure-lowering effects (clinic systolic blood pressure lower by 4 mm Hg [P=0.0112]; diastolic blood pressure lower by 3 mm Hg [P=0.0032]) in conjunction with a 94.5 dynes × sex/cm5 lower total peripheral resistance (P=0.0368). There were no significant changes observed to ambulatory blood pressure, arterial stiffness, heart rate variability, or cardiac output with the addition of empagliflozin. CONCLUSIONS: Adding sodium-glucose cotransporter 2 inhibitor treatment to angiotensin-converting enzyme inhibitor resulted in an expected GFR dip, suppression of oxidative stress markers, additive declines in blood pressure and total peripheral resistance. These changes are consistent with a protective physiologic profile characterized by the lowering of intraglomerular pressure and related cardiorenal risk when adding a sodium-glucose cotransporter 2 inhibitor to conservative therapy. REGISTRATION: URL: https://www. CLINICALTRIALS: gov; Unique identifier: NCT02632747.

Renal Hemodynamic Function and RAAS Activation Over the Natural History of Type 1 Diabetes.

RATIONALE & OBJECTIVE: The renin-angiotensin-aldosterone system (RAAS) is associated with renal and cardiovascular disease in diabetes. Unfortunately, early RAAS blockade in patients with type 1 diabetes mellitus (T1DM) does not prevent the development of complications. We sought to examine the role of hyperfiltration and RAAS activation across a wide range of T1DM duration to better understand renal hemodynamic status in patients with T1DM. STUDY DESIGN: Post hoc analysis of blood samples. SETTING & PARTICIPANTS: 148 Canadian patients with T1DM: 28 adolescents (aged 16.2±2.0 years), 54 young adults (25.4±5.6 years), and 66 older adults (65.7±7.5 years) studied in a clinical investigation unit. EXPOSURE: Angiotensin II infusion (1ng/kg/min; a measure of RAAS activation) during a euglycemic clamp. OUTCOMES: Glomerular filtration rate measured using inulin clearance, effective renal plasma flow measured using para-aminohippurate, afferent (RA) and efferent (RE) arteriolar resistances, and glomerular hydrostatic pressure estimated using the Gomez equations. RESULTS: In a stepwise fashion, glomerular filtration rate, effective renal plasma flow, and glomerular hydrostatic pressure were higher, while renal vascular resistance and RA were lower in adolescents versus young adults versus older adults. RE was similar in adolescents versus young adults but was higher in older adults. Angiotensin II resulted in blunted renal hemodynamic responses in older adults (renal vascular resistance increase of 3.3% ± 1.6% vs 4.9% ± 1.9% in adolescents; P<0.001), suggesting a state of enhanced RAAS activation. LIMITATIONS: Homogeneous study participants limit the generalizability of findings to other populations. Studying older adult participants with T1DM may be associated with a survivorship bias. CONCLUSIONS: A state of relatively low RAAS activity and predominant afferent dilation rather than efferent constriction characterize early adolescents and young adults with T1DM. This state of endogenous RAAS inactivity in early T1DM may explain why pharmacologic blockade of this neurohormonal system is often ineffective in reducing kidney disease progression in this setting. Older adults with long-standing T1DM who have predominant afferent constriction and RAAS activation may experience renoprotection from therapies that target the afferent arteriole. Further work is required to understand the potential role of non-RAAS pharmacologic agents that target RA in patients with early and long-standing T1DM.

Association between uric acid, renal haemodynamics and arterial stiffness over the natural history of type 1 diabetes.

AIMS: To examine the relationship between normal plasma uric acid (PUA) levels, renal haemodynamic function, arterial stiffness and plasma renin and aldosterone over a wide range of type 1 diabetes (T1D) durations in adolescents, young adults and older adults. MATERIALS AND METHODS: PUA, glomerular filtration rate (GFR), effective renal plasma flow (ERPF), vascular stiffness parameters (aortic augmentation index [AIx], carotid AIx, carotid femoral pulse wave velocity [cfPWV]), and plasma renin and aldosterone were measured during a euglycaemic clamp in people with T1D: 27 adolescents (mean ± SD age 16.8 ± 1.9 years), 52 young adults (mean ± SD age 25.6 ± 5.5 years) and 66 older adults (mean ± SD age 65.7 ± 7.5 years). RESULTS: PUA was highest in patients with the longest T1D duration: 197 ± 44 µmol/L in adolescents versus 264 ± 82 µmol/L in older adults (P < 0.001). Higher PUA correlated with lower GFR only in older adults, even after correcting for age, glycated haemoglobin and sex (ß = -2.12 ± 0.56; P = 0.0003), but not in adolescents or young adults. Higher PUA correlated with lower carotid AIx (ß = -1.90, P = 0.02) in adolescents. In contrast, PUA correlated with higher cfPWV (P = 0.02) and higher plasma renin (P = 0.01) in older adults with T1D. CONCLUSIONS: The relationship between higher PUA with lower GFR, increased arterial stiffness and renin angiotensin aldosterone system (RAAS) activation was observed only in older adults with longstanding T1D. T1D duration may modify the association between PUA, renal haemodynamic function and RAAS activation, leading to renal vasoconstriction and ischaemia. Further work must determine whether pharmacological PUA-lowering prevents or reverses injurious haemodynamic and neurohormonal sequelae of longstanding T1D, thereby improving clinical outcomes.

The relationships between markers of tubular injury and intrarenal haemodynamic function in adults with and without type 1 diabetes: Results from the Canadian Study of Longevity in Type 1 Diabetes.

OBJECTIVE: Our aim was to define the relationships between plasma biomarkers of kidney injury and intrarenal haemodynamic function (glomerular filtration rate [GFR], effective renal plasma flow [ERPF], renal vascular resistance [RVR]) in adults with type 1 diabetes (T1D). METHODS: The study sample comprised patients with longstanding T1D (duration ≥50 years), among whom 44 were diabetic kidney disease (DKD) resistors (eGFR >60 mL/min/1.73 m2 and <30 mg/d urine albumin excretion) and 22 had DKD, in addition to 73 control participants. GFRINULIN and ERPFPAH were measured, RVR was calculated, and afferent (RA )/efferent (RE ) areteriolar resistances were derived from Gomez equations. Plasma neutrophil gelatinase-associated lipocalin (NGAL), ß2 microglobulin (B2M), osteopontin (OPN) and uromodulin (UMOD) were measured using immunoassay kits from Meso Scale Discovery. RESULTS: Plasma NGAL, B2M and OPN were higher and UMOD was lower in DKD patients vs DKD resistors and non-diabetic controls. In participants with T1D, plasma NGAL inversely correlated with GFR (r = -0.33; P = 0.006) and ERPF (r = -0.34; P = 0.006), and correlated positively with RA (r = 0.26; P = 0.03) and RVR (r = 0.31; P = 0.01). In participants without T1D, NGAL and B2M inversely correlated with GFR (NGAL r = -0.18; P = 0.13 and B2M r = -0.49; P < 0.0001) and with ERPF (NGAL r = -0.19; P = 0.1 and B2M r = -0.42; P = 0.0003), and correlated positively with RA (NGAL r = 0.19; P = 0.10 and B2M r = 0.3; P = 0.01) and with RVR (NGAL r = 0.20; P = 0.09 and B2M r = 0.34; P = 0.003). Differences were significant after adjusting for age, sex, HbA1c, SBP and LDL. There were statistical interactions between T1D status, B2M and intrarenal haemodynamic function (P < 0.05). CONCLUSIONS: Elevated NGAL relates to intrarenal haemodynamic dysfunction in T1D, whereas elevated NGAL and B2M relate to intrarenal haemodynamic dysfunction in adults without T1D. These data may define a diabetes-specific interplay between tubular injury and intrarenal haemodynamic dysfunction.

Risk factors for diabetic kidney disease in adults with longstanding type 1 diabetes: results from the Canadian Study of Longevity in Diabetes.

Objectives: Diabetic kidney disease (DKD) is an independent predictor of cardiovascular morbidity and mortality in type 1 diabetes (T1D). We aimed to explore clinical and biochemical factors, including the achievement of American Diabetes Association (ADA) recommended targets associated with DKD in people living with T1D for ≥50 years. Methods: This was a post hoc analysis of a cross-sectional study of 75 participants enrolled in the Canadian Study of Longevity in T1D. We explored diabetes-related complications, including neuropathy, retinopathy, cardiovascular disease, and DKD. Study participants were dichotomized based on the achievement of ADA recommended targets as the low-target group (achieving ≤4 targets, n = 31) and high-target group (achieving >4 targets, n = 44). The outcome of interest was DKD defined by estimated glomerular filtration rate (eGFR) values <60/mL/min/1.73 m2 and/or 24-h albumin excretion >30 mg. Multivariable logistic regression models were employed to estimate odds ratios (ORs) for DKD with 95% confidence intervals (CIs). Results: Of the 75 participants with prolonged T1D duration (45% male, mean age 66 years), 25 participants had DKD and 50 did not. There was no statistical difference between the high- and low-target groups in terms of age and body mass index. eGFR was significantly higher and the prevalence of diabetic retinopathy was significantly lower in the high-target group. Older age at diagnosis of T1D and lower frequency component to high-frequency component ratio increased the odds of having DKD. Conclusions: In adults with prolonged T1D duration, older age at diagnosis and lower heart rate variability may be associated with DKD.

Dapagliflozin in focal segmental glomerulosclerosis: a combined human-rodent pilot study.

Focal segmental glomerulosclerosis (FSGS) is an important cause of nondiabetic chronic kidney disease (CKD). Sodium-glucose cotransporter 2 inhibition (SGLT2i) therapy attenuates the progression of diabetic nephropathy, but it remains unclear whether SGLT2i provides renoprotection in nondiabetic CKD such as FSGS. The primary aim of this pilot study was to determine the effect of 8 wk of dapagliflozin on glomerular filtration rate (GFR) in humans and in experimental FSGS. Secondary end points were related to changes in renal hemodynamic function, proteinuria, and blood pressure (BP). GFR (inulin) and renal plasma flow (para-aminohippurate), proteinuria, and BP were measured in patients with FSGS ( n = 10), and similar parameters were measured in subtotally nephrectomized (SNx) rats. In response to dapagliflozin, changes in GFR, renal plasma flow, and 24-h urine protein excretion were not statistically significant in humans or rats. Systolic BP (SBP) decreased in SNx rats (196 ± 26 vs. 165 ± 33 mmHg; P < 0.001), whereas changes were not statistically significant in humans (SBP 112.7 ± 8.5 to 112.8 ± 11.2 mmHg, diastolic BP 71.8 ± 6.5 to 69.6 ± 8.4 mmHg; P = not significant), although hematocrit increased (0.40 ± 0.05 to 0.42 ± 0.05%; P = 0.03). In archival kidney tissue from a separate patient cohort, renal parenchymal SGLT2 mRNA expression was decreased in individuals with FSGS compared with controls. Short-term treatment with the SGLT2i dapagliflozin did not modify renal hemodynamic function or attenuate proteinuria in humans or in experimental FSGS. This may be related to downregulation of renal SGLT2 expression. Studies examining the impact of SGLT2i on markers of kidney disease in patients with other causes of nondiabetic CKD are needed.

Renal hemodynamic effect of sodium-glucose cotransporter 2 inhibition in patients with type 1 diabetes mellitus.

BACKGROUND: The primary objective of this mechanistic open-label, stratified clinical trial was to determine the effect of 8 weeks' sodium glucose cotransporter 2 inhibition with empagliflozin 25 mg QD on renal hyperfiltration in subjects with type 1 diabetes mellitus (T1D). METHODS AND RESULTS: Inulin (glomerular filtration rate; GFR) and paraaminohippurate (effective renal plasma flow) clearances were measured in individuals stratified based on having hyperfiltration (T1D-H, GFR ≥ 135 mL/min/1.73m(2), n=27) or normal GFR (T1D-N, GFR 90-134 mL/min/1.73m(2), n=13) at baseline. Renal function and circulating levels of renin-angiotensin-aldosterone system mediators and NO were measured under clamped euglycemic (4-6 mmol/L) and hyperglycemic (9-11 mmol/L) conditions at baseline and end of treatment. During clamped euglycemia, hyperfiltration was attenuated by -33 mL/min/1.73m(2) with empagliflozin in T1D-H, (GFR 172±23-139±25 mL/min/1.73 m(2), P<0.01). This effect was accompanied by declines in plasma NO and effective renal plasma flow and an increase in renal vascular resistance (all P<0.01). Similar significant effects on GFR and renal function parameters were observed during clamped hyperglycemia. In T1D-N, GFR, other renal function parameters, and plasma NO were not altered by empagliflozin. Empagliflozin reduced hemoglobin A1c significantly in both groups, despite lower insulin doses in each group (P≤0.04). CONCLUSIONS: In conclusion, short-term treatment with the sodium glucose cotransporter 2 inhibitor empagliflozin attenuated renal hyperfiltration in subjects with T1D, likely by affecting tubular-glomerular feedback mechanisms. CLINICAL TRIAL REGISTRATION URL: http://www.clinicaltrials.gov. Unique identifier: NCT01392560.

Transdermal contraception and the renin-angiotensin-aldosterone system in premenopausal women.

The oral contraceptive pill (OCP) activates the renin-angiotensin-aldosterone system (RAAS) through first-pass hepatic metabolism. Although usually benign, RAAS activation may have detrimental effects on renal and hemodynamic function in some women. Since combined hormonal contraception with the transdermal patch (EVRA) does not undergo first-pass hepatic metabolism, we hypothesized that the RAAS response would be different from that of OCP subjects. Thirty-five nonsmoking, premenopausal women (15 control subjects, 10 OCP subjects, and 10 contraceptive patch subjects) without evidence of cardiovascular disease, renal disease, or diabetes were studied. Baseline angiotensinogen, renin, angiotensin II, aldosterone, and plasma renin activity were assessed along with hormonal and hemodynamic responses to simulated orthostatic stress using incremental lower body negative pressure (LBNP; -15, -25, and -40 mmHg). Baseline levels of angiotensinogen, angiotensin II, and plasma renin activity were significantly higher in OCP subjects compared with normotensive control and contraceptive patch subjects (P < 0.05), whereas aldosterone was significantly higher in OCP versus control subjects only (P < 0.05). Plasma renin levels were significantly lower at baseline in contraceptive patch subjects compared with normotensive control and OCP subjects (P < 0.05). In response to LBNP, increases in renin, angiotensin II, and aldosterone were attenuated in contraceptive patch subjects in conjunction with an exaggerated decline in mean arterial pressure (P < 0.05 vs. control and OCP subjects). The contraceptive patch in healthy premenopausal women is associated with an impaired ability to maintain blood pressure in response to LBNP, possibly due to insensitivity of the endogenous RAAS. Further evaluation may be beneficial in women with kidney disease.

The effect of sex on humanin levels in healthy adults and patients with uncomplicated type 1 diabetes mellitus.

Diabetes mellitus (DM) is associated with a loss of renal and vascular protection in women compared with men, but the responsible mechanisms are unclear. Recent experimental work implicated humanin (HN) as a novel cytoprotective hormone in DM. Our goal was to measure sex-related differences in HN levels in uncomplicated type 1 DM patients (T1D) and healthy controls (HC), as well as the interaction between HN, circulating neurohormones, and vascular function. Plasma HN, cGMP and aldosterone, blood pressure (BP), glomerular filtration rate, and effective renal plasma flow (inulin and para-aminohippurate) were measured in HC (11 men, 10 women) and T1D (23 men and 18 women) during clamped euglycemia (4-6 mmol·L(-1)). Plasma HN levels were generally lower in HC men by comparison with the women, but the differences were not statistically significant. In contrast, levels in the T1D men were higher compared with the T1D women (p = 0.026) and HC men (p < 0.0001). In the HC men, but not the women, HN correlated negatively with BP, but not with renal function, cGMP, or aldosterone. In the T1D men, HN negatively correlated with plasma cGMP. In the T1D women, HN did not correlate with neurohormones or vascular function. Future work should determine the role of HN in the pathogenesis of sex-related vascular function differences in DM.

A randomized cross-over comparison of short-term exposure of once-daily extended release tacrolimus and twice-daily tacrolimus on renal function in healthy volunteers.

Calcineurin inhibitor nephrotoxicity remains an issue for transplant recipients. The pharmacokinetic profile (PK) of the once-daily tacrolimus extended release (Tac-ER) includes equivalent exposure [AUC(0-24 h) ] but lower Cmax versus twice-daily tacrolimus immediate release (Tac-IR). We hypothesized that the unique PK profiles would result in pharmacodynamic differences in renal function. Nineteen healthy male subjects were allocated to once-daily Tac-ER and twice-daily Tac-IR in a prospective, randomized, two period, cross-over study. Tacrolimus was titrated to achieve trough levels of 8-12 ng/ml. Twenty four hours ERPF and GFR estimated by para-aminohippurate and sinistrin clearance were performed at baseline and at the end of each 10-day dosing period. Mean Tac C0 was 11.0 ± 2.2 and 11.3 ± 1.8 ng/ml for Tac-ER and Tac-IR, respectively. The mean Effective 24 h renal plasma flow (ERPF) was significantly higher with Tac-ER compared with Tac-IR (658 ± 127 vs. 610 ± 93 ml/min/1.73 m(2) , P = 0.046). There was a trend to a greater mean GFR over 24 h for Tac-ER at 114.5 ± 13.6 ml/min/1.73 m(2) compared with 108.9 ± 9.7 ml/min/1.73 m(2) for Tac-IR, P = 0.116. Under controlled physiological conditions, ERPF was significantly improved with Tac-ER compared with Tac-IR, likely owing to the differing PKs of these tacrolimus preparations (ClinicalTrials.gov Identifier: NCT01681134).

Urinary ACE2 in healthy adults and patients with uncomplicated type 1 diabetes.

Angiotensin-converting enzyme 2 (ACE2) is expressed in the kidney and may be renoprotective. We determined whether urinary ACE2 enzyme activity and protein levels (ELISA), as well as angiotensinogen and ACE, are elevated during clamped euglycemia (4-6 mmol·L(-1)) in patients with uncomplicated type 1 diabetes (T1D, n = 58) compared with normoglycemic controls (n = 21). We also measured the effect of clamped hyperglycemia (9-11 mmol·L(-1)) on each urinary factor in T1D patients. Urinary ACE2 activity and protein levels were higher during clamped euglycemia in T1D compared with the controls (p < 0.0001). In contrast, urinary angiotensinogen levels (p = 0.27) and ACE excretion (p = 0.68) did not differ. In response to clamped hyperglycemia in T1D, urinary ACE2 protein decreased (p < 0.0001), whereas urinary ACE2 activity as well as angiotensinogen and ACE levels remained unchanged. Urinary ACE2 activity and protein expression are increased in T1D patients prior to the onset of clinical complications. Further work is required to determine the functional role of urinary ACE2 in early T1D.

Long-term hemodynamic and molecular effects persist after discontinued renin-angiotensin system blockade in patients with type 1 diabetes mellitus.

Animal studies suggest temporary renin-angiotensin system (RAS) blockade enhances long-term vascular protective effects; however, this is not established in humans. Here we evaluated the long-term effects of prior RAS blockade on hemodynamic function, urinary measures of inflammation, and tissue antioxidant mRNA expression in patients with type 1 diabetes mellitus (T1DM) who participated in the 5-year Renin Angiotensin System Study (RASS). At 4 years after completing the RASS and discontinuing study medication, renal hemodynamic responses to clamped hyperglycemia were significantly greater in 18 patients in the RAS blockade group compared to 9 patients of the placebo-treated group. Individuals who had received RAS blockade also exhibited higher flow-mediated vasodilatation, reduced urinary cytokine excretion in response to hyperglycemia, and increased skin mRNA expression of superoxide dismutase-1 and catalase. Thus, patients with uncomplicated T1DM who received prior RAS blockade for 5 years maintain long-term effects on renal hemodynamic and systemic vascular function, inflammatory pathways in the kidney, and antioxidant enzyme expression in skin 4 years after discontinuation of therapy. Our findings suggest that sustained long-term protective effects of finite RAS inhibition requires further study.

Hyperfiltration and effect of nitric oxide inhibition on renal and endothelial function in humans with uncomplicated type 1 diabetes mellitus.

Studies of experimental diabetes mellitus (DM) suggest that increased nitric oxide (NO) bioactivity contributes to renal hyperfiltration. However, the role of NO in mediating hyperfiltration has not been fully elucidated in humans. Our aim was to examine the effect of NO synthase inhibition on renal and peripheral vascular function in normotensive subjects with uncomplicated type 1 DM. Renal function and brachial artery flow-mediated vasodilatation (FMD) were measured before and after an intravenous infusion of the NO synthase inhibitor N(G)-nitro-l-arginine methyl ester (l-NMMA) in 21 healthy control and 37 type 1 DM patients. Measurements in DM participants were made under clamped euglycemic conditions. The effect of l-NMMA on circulating and urinary NO metabolites (NO(x)) and cGMP and on urinary prostanoids was also determined. Baseline characteristics were similar in the two groups. For analysis, the DM patients were divided into those with hyperfiltration (DM-H, n = 18) and normal glomerular filtration rate (GFR) levels (DM-N, n = 19). Baseline urine NO(x) and cGMP were highest in DM-H. l-NMMA led to a decline in GFR in DM-H (152 ± 16 to 140 ± 11 ml·min(-1)·1.73 m(-2)) but not DM-N or healthy control participants. The decline in effective renal plasma flow in response to l-NMMA (806 ± 112 to 539 ± 80 ml·min(-1)·1.73 m(-2)) in DM-H was also exaggerated compared with the other groups (repeated measures ANOVA, P < 0.05), along with declines in urinary NO(x) metabolites and cGMP. Baseline FMD was lowest in DM-H compared with the other groups and did not change in response to l-NMMA. l-NMMA reduced FMD and plasma markers of NO bioactivity in the healthy control and DM-N groups. In patients with uncomplicated type 1 DM, renal hyperfiltration is associated with increased NO bioactivity in the kidney and reduced NO bioactivity in the systemic circulation, suggesting a paradoxical state of high renal and low systemic vascular NO bioactivity.

Systemic hemodynamic function in humans with type 1 diabetes treated with protein kinase Cß inhibition and renin-angiotensin system blockade: a pilot study.

The protein kinase Cß (PKCß) system has been implicated in the deleterious vascular responses to hyperglycemia and angiotensin II (Ang II) in experimental models of diabetes (DM). Whether these interactions are important in humans is unknown. Flow-mediated vasodilatation (FMD) was measured during clamped euglycemia and hyperglycemia, before and after randomization to PKCß inhibition (ruboxistaurin; RBX, 32 mg daily, n = 13) or a placebo (n = 7) for 8 weeks in renin-angiotensin system (RAS) blockade-treated subjects with type 1 DM. Blood pressure responses to infused Ang II were measured before and after randomization to RBX or a placebo. The RBX and placebo groups displayed similar clinical characteristics. Before RBX, FMD declined in response to hyperglycemia (6.8% ± 2.8% to 4.9% ± 1.8%). This effect was reversed after treatment with RBX (5.6% ± 3.1% to 6.0% ± 1.6% (within-group change, p = 0.009 (ANOVA)). No changes were observed in the placebo group. Infused Ang II was associated with hypertensive responses in the RBX and placebo groups (p < 0.05 (ANOVA)), and RBX did not influence this effect. In conclusion, RBX blunted the effect of hyperglycemia on FMD, suggesting that PKCß may modulate endothelial function in type 1 DM. The lack of effect on Ang II responses suggests that PKCß inhibition may act through non-RAS pathways in humans with DM.

Vascular dysfunction in women with a history of preeclampsia and intrauterine growth restriction: insights into future vascular risk.

BACKGROUND: Women with a history of placental disease are at increased risk for the future development of vascular disease. It is unknown whether preexisting endothelial dysfunction underlies both the predisposition to placental disease and the later development of vascular disease. The aim of this study was to assess vascular function in postpartum women and to determine whether differences emerged depending on the presentation of placental disease. METHODS AND RESULTS: Women with a history of early-onset preeclampsia (n=15), late-onset preeclampsia (n=9), intrauterine growth restriction without preeclampsia (n=9), and prior normal pregnancy (n=16) were studied 6 to 24 months postpartum. Flow-mediated vasodilatation and flow-independent (glyceryl trinitrate-induced) vasodilatation were studied through the use of high-resolution vascular ultrasound examination of the brachial artery. Arterial stiffness was assessed by pulse-wave analysis (augmentation index). Laboratory assessment included circulating angiogenic factors (vascular endothelial growth factor, soluble fms-like tyrosine kinase 1, placental growth factor, and soluble endoglin). Flow-mediated vasodilatation was significantly reduced in women with previous early-onset preeclampsia and intrauterine growth restriction compared with women with previous late-onset preeclampsia and control subjects (3.2±2.7% and 2.1±1.2% versus 7.9±3.8% and 9.1±3.5%, respectively; P<0.0001). Flow-independent vasodilatation was similar among all groups. Similarly, the radial augmentation index was significantly increased among women with previous early-onset preeclampsia and intrauterine growth restriction, but not among late preeclamptic women and control subjects (P=0.0105). Circulating angiogenic factors were similar in all groups. CONCLUSION: Only women with a history of early-onset preeclampsia or intrauterine growth restriction without preeclampsia exhibit impaired vascular function, which might explain their predisposition to placental disease and their higher risk of future vascular disease.

Vasopressin associated with renal vascular resistance in adults with longstanding type 1 diabetes with and without diabetic kidney disease.

OBJECTIVE: Arginine vasopressin (AVP) and its surrogate, copeptin, have been implicated in diabetic kidney disease (DKD) pathogenesis, which develops in a subset of people with longstanding type 1 diabetes, but not in others (DKD Resistors). We hypothesized that patients with DKD would exhibit higher copeptin concentrations vs. DKD Resistors. METHODS: Participants with type 1 diabetes (n = 62, duration ≥50 years) were stratified into 42 DKD Resistors and 20 with DKD (eGFR ≤60 mL/min/1.73m2 or ≥30 mg/day urine albumin), and age/sex-matched controls (HC, n = 74) were included. Glomerular filtration rate (GFR) and effective renal plasma flow (ERPF) were calculated by inulin and p-aminohippurate clearance before and after angiotensin II (ang II) infusion. Renal vascular resistance (RVR) was calculated as mean arterial pressure/renal blood flow. Plasma copeptin, renin, aldosterone, neutrophil gelatinase-associated lipocalin (NGAL), and urea concentrations were measured, along with 24-h urine volume. RESULTS: DKD resistors had lower copeptin (95% CI: 4.0 [3.4-4.8] pmol/l) compared to DKD (5.8 [4.5-7.6] pmol/l, p = 0.02) and HC (4.8 [4.1-5.5] pmol/l, p = 0.01) adjusting for age, sex and HbA1c. In type 1 diabetes, higher copeptin correlated with lower GFR (r: -0.32, p = 0.01) and higher renin concentration (r: 0.40, p = 0.002) after multivariable adjustments. These relationships were not evident in HC. Copeptin inversely associated with RVR change following exogenous ang II only in participants with type 1 diabetes (ß ±â€¯SE: -6.9 ±â€¯3.4, p = 0.04). CONCLUSIONS: In longstanding type 1 diabetes, copeptin was associated with intrarenal renin-angiotensin-aldosterone system (RAAS) activation and renal hemodynamic function, suggesting interplay between AVP and RAAS in DKD pathogenesis.

Age is a determinant of acute hemodynamic responses to hyperglycemia and angiotensin II in humans with uncomplicated type 1 diabetes mellitus.

Hyperglycemia is associated with hemodynamic changes in type 1 diabetes (DM), acting in part through renin-angiotensin system activation. Since aging is associated with vascular dysfunction in DM, we hypothesized that acute hemodynamic responses to clamped hyperglycemia and infused ANG II would be exaggerated in older adults compared with a group of adolescent/young adults with type 1 DM. Renal hemodynamic function, blood pressure, and arterial stiffness were assessed in adolescent/young adults (n = 34; mean age: 18 +/- 3 yr) and older adults (n = 32; mean age: 45 +/- 9 yr). Studies were performed during clamped euglycemia (4-6 mmol/l) and hyperglycemia (9-11 mmol/l). Renal and systemic hemodynamic responses to ANG II were measured during clamped euglycemia in diabetic subjects. ANG II responses were also assessed in a cohort of non-DM subjects (n = 97; mean age: 26; age range: 18-40 yr). Older DM adults exhibited higher baseline blood pressure, arterial stiffness, and renal vascular resistance, and lower glomerular filtration rate (GFR) and effective renal plasma flow, compared with adolescent/young DM adults (P < 0.05). Clamped hyperglycemia was associated with exaggerated peripheral and renal hemodynamic responses uniquely in older DM adults; only GFR increased in adolescent/young DM adults. ANG II infusion also produced exaggerated vasoconstrictive responses in older DM adults vs. adolescent/young DM adults (P < 0.05). The independent effect of age on hemodynamic responses to hyperglycemia and ANG II was confirmed using multivariate regression analysis in DM subjects (P < 0.05), and results were still significant when participants were matched for DM duration. Age-related alterations in hemodynamic function and ANG II response were not observed in healthy non-DM control subjects. Acute hemodynamic responses to clamped hyperglycemia and ANG II were exaggerated in older subjects with type 1 DM, highlighting an important interaction between age and factors that contribute to the pathogenesis of acute vascular dysfunction in DM.

The angiotensin II receptor type 2 polymorphism influences haemodynamic function and circulating RAS mediators in normotensive humans.

BACKGROUND: The haemodynamic responses to angiotensin II type 1 (AT1) receptor blockade may be mediated in part by interactions between angiotensin II and the angiotensin II type 2 receptor (AT2R). An AT2R G1675A gene polymorphism has been described, but the functional effects of this polymorphism are unknown. METHODS: Haemodynamic function, circulating renin-angiotensin system mediators and norepinephrine were measured in young healthy subjects at baseline and at 2 and 4 weeks after treatment with irbesartan. Subjects were divided into two groups on the basis of the AT2R G1675A gene polymorphism: GG subjects (n = 12) and AA/GA subjects (n = 22). RESULTS: AA/AG subjects exhibited hypotensive and renal vasodilatory responses to irbesartan at 4 weeks, but GG subjects did not. In accord with haemodynamic effects, circulating aldosterone levels were suppressed in AA/AG, while circulating norepinephrine levels were augmented only in GG subjects. In contrast, increases in circulating renin, angiotensin II and plasma renin activity after irbesartan were exaggerated in AA/AG subjects. CONCLUSIONS: The AT2R G1675A polymorphism is a determinant of haemodynamic responses to AT1 receptor blockade, an effect that may be due to influences on aldosterone escape.

Endothelial nitric oxide synthase gene polymorphisms and the renal hemodynamic response to L-arginine.

Nitric oxide is generated from L-arginine by nitric oxide synthase (NOS), an enzyme that exists in several isoforms. Some studies found that a polymorphism (G894T) in the endothelial NOS gene was associated with decreased nitric oxide bioactivity and vascular complications. However, it is not known whether the enzyme had a reduced activity. Here we measured the effect of an infusion of L-arginine on renal hemodynamic function in subjects segregated by the presence or absence of the T allele. If this polymorphism represented a functional variant, subjects with the GT/TT form should exhibit a blunted renal hemodynamic response to L-arginine compared to those with a GG allele. All subjects were given a diet controlled for sodium and protein intake. GG subjects had lower mean arterial pressure and an augmented glomerular filtration rate at baseline. In response to a graded L-arginine infusion, this group had significant changes in effective renal plasma flow, glomerular filtration rate, filtration fraction, renal vascular resistance, and renal blood flow. The renal response to L-arginine in GT/TT subjects was blunted. Circulating cGMP levels and endothelial NOS mRNA expression, measured in skin biopsies by real-time PCR, did not differ between the groups. Our study shows that the G894T allele of endothelial NOS is associated with a blunted response to L-arginine, suggesting this polymorphism may be a functional variant in humans.

Estimating GFR by Serum Creatinine, Cystatin C, and ß2-Microglobulin in Older Adults: Results From the Canadian Study of Longevity in Type 1 Diabetes.

INTRODUCTION: Glomerular filtration rate (GFR) is routinely used for clinical assessment of kidney function. However, the accuracy of estimating equations in older adults is uncertain. METHODS: In 66 adults with ≥50 years type 1 diabetes (T1D) duration and 73 nondiabetic controls from age/sex-matched subgroups (65 ± 8 years old and 77[55%] were women) we evaluated the performance of estimated GFR (eGFR) by creatinine (Modification of Diet and Renal Disease [MDRD], Chronic Kidney Disease-Epidemiology [CKD-EPI]cr), cystatin C (CKD-EPIcys, CKD-EPIcr-cys), and ß2-microglobulin (ß2M) compared with measured GFR by inulin clearance (mGFR). Performance was evaluated using metrics of bias (mean difference), precision (SD), and accuracy (proportion of eGFR that differed by >20% of mGFR). RESULTS: Mean mGFR was 104 ± 18 ml/min per 1.73 m2 (range: 70-154 ml/min per 1.73 m2) and was not different between T1D and controls (103 ± 17 vs. 105 ± 19 ml/min per 1.73 m2, P = 0.39). All equations significantly underestimated mGFR (bias: -15 to -30 ml/min per 1.73 m2, P < 0.001 for all comparisons) except for ß2M, which had bias of 1.9 ml/min per 1.73 m2 (P = 0.61). Bias was greatest in cystatin C-based equations. Precision was lowest for ß2M (SD: 43.5 ml/min per 1.73 m2, P < 0.001 for each comparison). Accuracy was lowest for CKD-EPIcysC (69.1%, P < 0.001 for each comparison). Cystatin C-based equations demonstrated greater bias and lower accuracy in older age subgroups (<60, 60-69, ≥70 years). All equations demonstrated greater bias across higher ranges of mGFR (60-89, 90-119, ≥120 ml/min per 1.73 m2). Results were similar between T1D and controls except that ß2M had lower performance in T1D. CONCLUSION: Better estimates of GFR in older adults are needed for research and clinical practice, as this subgroup of the population has an amplified risk for the development of chronic kidney disease (CKD) that requires accurate GFR estimation methods.