Vascular acetylcholine response during chronic NO synthase inhibition: in vivo versus in vitro

Objective: The aim of this study was to compare the response to NO-mediated vasodilators in vivo and in vitro during chronic NO synthase inhibition. Methods:NG-Nitro-l-arginine-methyl ester (l-NAME, 0.4 g/l) or vehicle was administered in the drinking water for 6 weeks to male Wistar rats weighing 220-240 g. The effect of acetylcholine and sodium nitroprusside was examined in vivo, on systemic blood pressure and heart rate and in vitro, on the precontracted isolated mesenteric artery. The in vivo response to both vasodilators was examined in awake rats monitored by an indwelling catheter in the femoral artery. Isolated segments of the third-generation mesenteric artery were examined in vitro with a Mulvany dual myograph after precontraction with noradrenaline. Results: In isolated mesenteric arteries obtained from rats chronically treated with l-NAME, the initial relaxant response to acetylcholine was significantly decreased whereas that to sodium nitroprusside was enhanced. A late acetylcholine-induced contractile response was present and abolished by indomethacin. In vivo, the hypotensive action of sodium nitroprusside was also enhanced in the l-NAME-treated rats. Acetylcholine reduced blood pressure in the l-NAME-treated hypertensive animals more than in normotensive controls, but less than in control rats infused intravenously with noradrenaline at a dose increasing their blood pressure to hypertensive levels. Conclusions: The NO-mediated vasodilation induced by acetylcholine is attenuated during chronic NO synthase inhibition, both in vivo and in vitro. The blunted hypotensive response to acetylcholine can be demonstrated only if blood pressure of control rats is acutely increased to hypertensive level

P-621: Metabolic determinants of proximal sodium reabsorption in healthy women

Increased reabsorption of sodium (Na) in the proximal segments of the nephron is present in hypertensive patients. Studies performed in men reported an association between proximal Na reabsorption and features of the metabolic syndrome. This study explores the relationship between metabolic variables and proximal Na reabsorption in healthy women. This study is population-based and cross-sectional examining 661 healthy women aged 40-75y. After 15 minutes rest, blood pressure was measured 3 times, blood was drawn and urine was collected. Fractional excretion of endogenous lithium (FELi) was used as an indirect marker of proximal sodium reabsorption, lithium being only reabsorbed in the proximal tubule. All women receiving blood pressure, blood glucose or lipid lowering therapy were excluded. Correlations: FELi was positively correlated with the fractional excretion of sodium (FENa, r=0.3, p<0.001). FELi was negatively associated with total cholesterol (r=-0.14, P<0.0001), LDL-cholesterol (r=-0.16, P<0.0001), BMI (r=-0.08, P<0.05) and weight (r=-0.09, P<0.05). Menopausal status or a family history of hypertension did not affect the associations. Simple linear regression analysis: age, waist circumference, waist/hip ratio, systolic blood pressure, diastolic blood pressure, Hdl cholesterol, triglycerides, serum uric acid or a family history of hypertension were not significant predictors of FELi. BMI was a significant predictor but the strongest relationships were found between FELi and total cholesterol, LDL-cholesterol and FENa. Multivariate linear regression model: When significant predictors of FELi were examined in a multivariate linear regression model also controlling for age, weight, systolic blood pressure and FENa, total cholesterol (p=0.003) or LDL-cholesterol (p=0.001) significantly and independently predicted FELi. In conclusion, these data suggest that metabolic parameters, in particular total cholesterol and LDL-cholesterol and to a lesser extent weight and BMI, are associated with increased proximal Na reabsorption in a healthy untreated women population. Considering the possible link between increased reabsorption of sodium and the development of hypertension, a major cardiovascular risk factor, this association may provide an additional hypothesis for the increased cardiovascular risk of subjects with the metabolic syndrom

P-537: Pioglitazone blunts the blood pressure response to angiotensin II in insulin-resistant zucker rats

Thiazolidinediones are high-affinity agonists of peroxisome proliferator-activated receptors (PPAR)-γ which have been found to lower blood pressure (BP) in animal models of diabetes and reno-vascular hypertension. The mechanisms leading to this decrease in blood pressure are still unclear. Since the renin-angiotensin system is a main regulator of blood pressure, we investigated the effects of pioglitazone on blood pressure, plasma renin activity and on the blood pressure response to exogenous angiotensin II in insulin-resistant Zucker rats. Pioglitazone 20mg/kg/d or vehicle were administered for 4 weeks to 8-weeks old fa/fa Zucker rats. Pioglitazone-treated rats were heavier than vehicle-treated rats (respectively 481g±8g vs 437±5g, p=0.0002, mean±SEM) and ate more (35.6±0.5g/d vs 28.9±0.3, p<0.0001). The increase in blood sugar after an intra-venous glucose tolerance test was significantly attenuated in the pioglitazone treated rats at 10,15 and 30 minutes. Systolic (SBP), diastolic (DBP) blood pressure and heart rate (HR) were lower in pioglitazone-treated rats: SBP: 124±3 mmHg vs 144±3, p<0.001; DBP: 80±2 vs 94±2, p<0.001; HR: 369±6 vs 397±8, p<0.01. The BP response to exogenous angiotensin II was significantly attenuated in pioglitazone treated rats. With the 25 ng/kg Ang II dose the increase in BP was 27.8±2.4 mmHg with pioglitazone and 37.5±3.3 with the vehicle (p=0.04) and with the Ang II 100 ng/kg dose the increase in BP was respectively 36.1±2.7 mmHg and 49.2±2.3 (p=0.003). Plasma renin activity was comparable in both groups. In conclusion, these results show that pioglitazone blunts the BP response to angiotensin II in insulin-resistant Zucker rats. This effect may partially explain the blood pressure lowering effect of PPAR-γ agonist

P-620: Pioglitazone stimulates renin and favors sodium retention and weight gain in healthy subjects

Glitazones induce peripheral edema through an unknown mechanism in up to 20% of cases. This study examines the effects of pioglitazone (PIO) on renal sodium handling and renal hemodynamics in healthy male volunteers (HV) exposed to a high (HS) and low (LS) sodium diet. The influence of PIO on plasma renin activity (PRA), aldosterone and atrial natriuretic peptide (ANP) was examined. Nine HV aged 22-28 y were enrolled. BMI, blood pressure and glucose tolerance were normal. The study had a double-blind, randomized, placebo controlled, twofold cross-over design. Each subject received either PIO 45 mg qd or placebo qd, for 6 weeks, with 2 weeks wash-out. From weeks 1-4, subjects were on their usual diet. During weeks 5 and 6, subjects were either on a LS or a HS diet for a week which was followed by ambulatory blood pressure measurements, hormonal measurements and renal function studies. The differences between PIO and placebo effects were examined (median, range among all subjects). No subject developed edema. Insulin sensitivity, systolic and diastolic blood pressure, glomerular filtration rate, renal plasma flow or filtration fraction did not change significantly with PIO. Weight increased with PIO in 7/9 subjects while on a LS diet (0.7kg; -1-2.9) and in 6/9 while on a HS diet (1.1kg; -1.5-3.4). Median sodium (Na) excretion with placebo was of 21.2mmol/24h and 239.7mmol/24h respectively while on a LS and HS diet. Urinary Na excretion decreased with PIO in 6/9 subjects on a LS diet (-12.2mmol/24h; -21-8.7, p=0.05), and in 5/9 subjects on a HS diet (-30mmol/24h; -344-69). Na clearance decreased in 6/9 subjects on a LS diet (-0.05 ml/min; -0.11-0.05) and in 6/9 subjects on a HS diet (-0.15 mmol/ml; -2.7-0.4). Na clearance at the proximal level decreased with PIO in 8/9 subjects on a LS diet (-4.9 ml/min; -14.4-1.5, p=0.01) and in 5/9 subjects on a HS diet (-2 ml/min; -43-8.5). PRA increased with PIO in 8/9 subjects on a LS diet (0.16 ng/ml/h; -0.07-0.8, p=0.02) and on a HS diet (0.09 ng/ml/h; -0.1-0.21, p=0.03). Aldosterone increased in 5/9 subjects on a LS diet (6 pg/ml; -19-144) and in 6/9 subjects on a HS diet (5 pg/ml; -20-74). ANP levels did not change. In conclusion, pioglitazone increases PRA, independently from Na intake and favors Na retention in healthy volunteers. This mechanism could contribute to the development of edema in subjects treated with glitazone

Blood pressure and vascular determinants of glomerular filtration rate decline in diabetic kidney disease

ObjectiveIn patients with type 2 diabetes and diabetic kidney disease (DKD), explore the relationship between estimated glomerular filtration rate decline (eGFR-d) and simultaneously assessed vascular risk markers including office, ambulatory or central blood pressure, pulse pressure, carotid-femoral pulse wave velocity (PWV), carotid intima-media thickness (IMT) and renal resistive indexes (RRI).Research design and methodsAt baseline, vascular risk markers were measured in addition to the routine clinical workup. The eGFR-d was based on 2000–2019 creatinine values. Parameters were compared by eGFR-d quartiles. Regression models of eGFR-d and vascular markers were assessed.ResultsIn total, 135 patients were included. Mean age was 63.8 ± 10.8y, baseline eGFR 60.2 ± 26.4 ml/min/1.73 m2 and urine albumin-creatinine ratio (ACR) 49 ± 108 mg/mmol. Mean eGFR-d was based on 43 ± 39 creatinine values within a time span of 7.0 ± 1.9y. The average yearly eGFR decline was −1.8 ± 3.0 ml/min/1.73 m2 ranging from −5.8 ± 2.3 in the first quartile to +1.4 ± 1.7 in the fourth quartile. Mean 24 h systolic (SBP) and diastolic (DBP) blood pressure were 126 ± 17 and 74 ± 9 mmHg. Mean PWV was 11.8 ± 2.8 m/s, RRI 0.76 ± 0.07 and IMT 0.77 ± 0.21 mm. SBP and pulse pressure correlated with eGFR-d but not DBP. 24 h SBP stood out as a stronger predictor of eGFR-d than office or central SBP. PWV and RRI correlated with eGFR decline in univariate, but not multivariate regression models including 24 SBP and ACR.ConclusionsIn this study, eGFR decline was highly variable in patients with type 2 diabetes and DKD. Twenty-four hour SBP provided an added value to the routine measurement of ACR in predicting eGFR decline, whereas PWV and RRI did not

Evidence against a direct role of klotho in insulin resistance

The klotho gene may be involved in the aging process. Klotho is a coactivator of FGF23, a regulator of phosphate and vitamin D metabolism. It has also been reported to be downregulated in insulin resistance syndromes and paradoxically to directly inhibit IGF-1 and insulin signaling. Our aim was to study klotho's regulation and effects on insulin and IGF-1 signaling to unravel this paradox. We studied klotho tissue distribution and expression by quantitative real-time polymerase chain reaction and Western blotting in obese Zucker rats and high-fat fed Wistar rats, two models of insulin resistance. Klotho was expressed in kidneys but at much lower levels (<1.5%) in liver, muscle, brain, and adipose tissue. There were no significant differences between insulin resistant and control animals. We next produced human recombinant soluble klotho protein (KLEC) and studied its effects on insulin and IGF-1 signaling in cultured cells. In HEK293 cells, FGF23 signaling (judged by FRS2-α and ERK1/2 phosphorylation) was activated by conditioned media from KLEC-producing cells (CM-KLEC); however, IGF-1 signaling was unaffected. CM-KLEC did not inhibit IGF-1 and insulin signaling in L6 and Hep G2 cells, as judged by Akt and ERK1/2 phosphorylation. We conclude that decreased klotho expression is not a general feature of rodent models of insulin resistance. Further, the soluble klotho protein does not inhibit IGF-1 and/or insulin signaling in HEK293, L6, and HepG2 cells, arguing against a direct role of klotho in insulin signaling. However, the hypothesis that klotho indirectly regulates insulin sensitivity via FGF23 activation remains to be investigate

Diabetic kidney disease in type 2 diabetes: a consensus statement from the Swiss Societies of Diabetes and Nephrology.

Diabetic kidney disease is highly prevalent in patients with type 2 diabetes and is a major cause of end-stage renal disease in Switzerland. Patients with diabetic kidney disease are among the most complex patients in diabetes care. They require a multifactorial and multidisciplinary approach with the goal to slow the decline in glomerular filtration rate (GFR) and cardiovascular morbidity. With this consensus we propose an evidence-based guidance to health care providers involved in the care of type 2 diabetic patients with diabetic kidney disease.First, there is a need to increase physician awareness and improve screening for diabetic kidney disease as early intervention may improve clinical outcomes and the financial burden. Evaluation of estimated GFR (eGFR) and spot urine albumin/creatinine ratio is recommended at least annually. Once it is diagnosed, glucose control and optimisation of blood pressure control with renin-angiotensin system blockers have been recommended as mainstay management of diabetic kidney disease for more than 20 years. Recent, high quality randomised controlled trials have shown that sodium-glucose cotransporter-2 (SGLT2) inhibition slows eGFR decline and cardiovascular events beyond glucose control. Likewise, mineralocorticoid receptor antagonism with finerenone has cardiorenal protective effects in diabetic kidney disease. Glucagon-like peptide-1 (GLP1) receptor agonists improve weight loss if needed, and decrease albuminuria and cardiovascular morbidity. Lipid control is also important to decrease cardiovascular events. All these therapies are included in the treatment algorithms proposed in this consensus. With advancing kidney failure, other challenges may rise, such as hyperkalaemia, anaemia and metabolic acidosis, as well as chronic kidney disease-mineral and bone disorder. These different topics and treatment strategies are discussed in this consensus. Finally, an update on diabetes management in renal replacement therapy such as haemodialysis, peritoneal dialysis and renal transplantation is provided. With the recent developments of efficient therapies for diabetic kidney disease, it has become evident that a consensus document is necessary. We are optimistic that it will significantly contribute to a high-quality care for patients with diabetic kidney disease in Switzerland in the future

The differential impact of a 6-versus 12-month pharmacist-led interprofessional medication adherence program on medication adherence in patients with diabetic kidney disease: the randomized PANDIA-IRIS study

Background: For every 100 patients with diabetes, 40 will develop diabetic kidney disease (DKD) over time. This diabetes complication may be partly due to poor adherence to their prescribed medications. In this study, we aimed to evaluate the differential impact of a 6- versus 12-month pharmacist-led interprofessional medication adherence program (IMAP) on the components of adherence (i.e., implementation and discontinuation) in patients with DKD, during and after the intervention.Methods: All included patients benefited from the IMAP, which consists in face-to-face regular motivational interviews between the patient and the pharmacist based on the adherence feedback from electronic monitors (EMs), in which the prescribed treatments were delivered. Adherence reports were available to prescribers during the intervention period. Patients were randomized 1:1 into two parallel arms: a 12-month IMAP intervention in group A versus a 6-month intervention in group B. Adherence was monitored continuously for 24 months post-inclusion during the consecutive intervention and follow-up phases. In the follow-up phase post-intervention, EM data were blinded. Blood pressure was measured by the pharmacist at each visit. The repeated measures of daily patient medication intake outcomes (1/0) to antidiabetics, antihypertensive drugs, and statins were modeled longitudinally using the generalized estimated equation in both groups and in both the intervention and the follow-up phases.Results: EM data of 72 patients were analyzed (34 in group A and 38 in group B). Patient implementation to antidiabetics and antihypertensive drugs increased during the IMAP intervention phase and decreased progressively during the follow-up period. At 12 months, implementation to antidiabetics was statistically higher in group A versus group B (93.8% versus 86.8%; Δ 7.0%, 95% CI: 5.7%; 8.3%); implementation to antihypertensive drugs was also higher in group A versus B (97.9% versus 92.1%; Δ 5.8%, 95% CI: 4.8%; 6.7%). At 24 months, implementation to antidiabetics and antihypertensive drugs remained higher in group A versus B (for antidiabetics: 88.6% versus 85.6%; Δ 3.0%, 95% CI: 1.7%; 4.4% and for antihypertensive drugs: 94.4% versus 85.9%; Δ 8.5%, 95% CI: 6.6%; 10.7%). No difference in pharmacy-based blood pressure was observed between groups. Implementation to statins was comparable at each time point between groups. Three patients discontinued at least one treatment; they were all in group B. In total, 46% (16/35) of patients in the 12-month intervention versus 37% (14/38) of patients in the 6-month intervention left the study during the intervention phase, mainly due to personal reasons.Conclusion: The IMAP improves adherence to chronic medications in patients with DKD. The longer the patients benefit from the intervention, the more the implementation increases over time, and the more the effect lasts after the end of the intervention. These data suggest that a 12-month rather than a 6-month program should be provided as a standard of care to support medication adherence in this population. The impact on clinical outcomes needs to be demonstrated.Clinical Trial Registration:Clinicaltrials.gov, identifier NCT04190251_PANDIA IRIS