Deleterious vascular effects of indoxyl sulfate and reversal by oral adsorbent AST-120.

BACKGROUND: In chronic kidney disease (CKD), blood vessels are permanently exposed to uremic toxins such as indoxyl sulfate (IS). We hypothesized that IS could alter vascular tone and that reducing its serum concentration could be beneficial. DESIGN: We studied acute and longer-term effects of IS and AST-120, an oral charcoal adsorbent, on vascular reactivity, endothelium integrity and expression of adhesion molecules VCAM-1 and ICAM-1 in aortic rings of normal and uremic wild type (WT) mice in vitro, and the cardiovascular effects of AST-120 in both WT and apoE-/- mice with CKD in vivo. RESULTS: In vitro, 1.0 mM IS acutely reduced vascular relaxation (64% for IS 1.0 mM vs. 80% for control, p < 0.05). The effect was more marked after 4 days exposure (39% for IS 1.0 mM 4 days; p < 0.001, prolonged vs. acute exposure), and was associated with endothelial cell loss and upregulation of ICAM-1/VCAM-1 expression. In vitro, AST-120 restored normal vascular function and prevented IS induced endothelial cell loss and ICAM-1/VCAM-1 upregulation. In vivo, AST-120 treatment of CKD mice (1) improved vascular relaxation (72% vs. 48% maximal relaxation in treated vs. untreated mice, p < 0.001), (2) reduced aortic VCAM-1 and ICAM-1 expression, (3) decreased aorta systolic expansion rate (9 ± 3% CKD vs. 14 ± 3% CKD + AST-120, p < 0.02), and (4) prevented the increase in pulse wave velocity (3.56 ± 0.17 m/s CKD vs. 3.10 ± 0.08 m/s CKD + AST-120, p < 0.006). Similar changes were observed in apoE-/- mice. CONCLUSION: IS appears to be an important contributor to the vascular dysfunction associated with CKD. AST-120 treatment ameliorates this dysfunction, possibly via a decrease in serum IS concentration.

Peritoneal delivery of sodium pyrophosphate blocks the progression of pre-existing vascular calcification in uremic apolipoprotein-E knockout mice.

Chronic kidney disease (CKD) is generally associated with disturbances of mineral and bone metabolism. They contribute to the development of vascular calcification (VC), a strong, independent predictor of cardiovascular risk. Pyrophosphate (PPi), an endogenous inhibitor of hydroxyapatite formation, has been shown to slow the progression of VC in uremic animals. Since in patients with CKD treatment is usually initiated for already existing calcifications, we aimed to compare the efficacy of PPi therapy with that of the phosphate binder sevelamer, using a uremic apolipoprotein-E knockout mouse model with advanced VCs. After CKD creation or sham surgery, 12-week-old female mice were randomized to one sham group and four CKD groups (n = 18-19/group). Treatment was initiated 8 weeks after left nephrectomy allowing prior VC development. Uremic groups received either intraperitoneal PPi (high dose, 1.65 mg/kg or low dose, 0.33 mg/kg per day), oral sevelamer (3 % in diet), or placebo treatment for 8 weeks. Both intima and media calcifications worsened with time in placebo-treated CKD mice, based on both quantitative image analysis and biochemical measurements. Progression of calcification between 8 and 16 weeks was entirely halted by PPi treatment, as it was by sevelamer treatment. PPi did not induce consistent bone histomorphometry changes. Finally, the beneficial vascular action of PPi probably involved mechanisms different from that of sevelamer. Further studies are needed to gain more precise insight into underlying mechanisms and to see whether PPi administration may also be useful in patients with CKD and VC.

Effects of pyrophosphate delivery in a peritoneal dialysis solution on bone tissue of apolipoprotein-E knockout mice with chronic kidney disease.

Vascular calcification (VC) is a risk factor for cardiovascular mortality in the setting of chronic kidney disease (CKD). Pyrophosphate (PPi), an endogenous molecule that inhibits hydroxyapatite crystal formation, has been shown to prevent the development of VC in animal models of CKD. However, the possibility of harmful effects of exogenous administration of PPi on bone requires further investigation. To this end, we examined by histomorphometry the bone of CKD mice after intraperitoneal PPi administration. After CKD creation or sham surgery, 10-week-old female apolipoprotein-E knockout (apoE(-/-)) mice were randomized to one non-CKD group or 4 CKD groups (n = 10-35/group) treated with placebo or three distinct doses of PPi, and fed with standard diet. Eight weeks later, the animals were killed. Serum and femurs were sampled. Femurs were processed for bone histomorphometry. Placebo-treated CKD mice had significantly higher values of osteoid volume, osteoid surface and bone formation rate than sham-placebo mice with normal renal function. Slightly higher osteoid values were observed in CKD mice in response to very low PPi dose (OV/BV, O.Th and ObS/BS) and, for one parameter measured, to high PPi dose (O.Th), compared to placebo-treated CKD mice. Treatment with PPi did not modify any other structural parameters. Mineral apposition rates, and other parameters of bone formation and resorption were not significantly different among the treated animal groups or control CKD placebo group. In conclusion, PPi does not appear to be deleterious to bone tissue in apoE(-/-) mice with CKD, although a possible stimulatory PPi effect on osteoid formation may be worth further investigation.

Possible involvement of microRNAs in vascular damage in experimental chronic kidney disease.

Chronic kidney disease (CKD) is associated with vascular calcifications and atherosclerosis. There is a need for novel predictors to allow earlier diagnosis of these disorders, predict disease progression, and improve assessment of treatment response. We focused on microRNAs since they are implicated in a variety of cellular functions in cardiovascular pathology. We examined changes of microRNA expression in aortas of CKD and non-CKD wild type mice and apolipoprotein E knock-out mice, respectively. Both vascular smooth muscle-specific miR-143 and miR-145 expressions were decreased in states of atherosclerosis and/or CKD or both, and the expression level of protein target Myocardin was increased. The inflammatory miR-223 was increased in more advanced stages of CKD, and specific protein targets NFI-A and GLUT-4 were dramatically decreased. Expression of miR-126 was markedly increased and expression of protein targets VCAM-1 and SDF-1 was altered during the course of CKD. The drug sevelamer, commonly used in CKD, corrected partially these changes in microRNA expression, suggesting a direct link between the observed microRNA alterations and uremic vascular toxicity. Finally, miR-126, -143 and -223 expression levels were deregulated in murine serum during the course of experimental CKD. In conclusion, these miRNAs could have role(s) in CKD vascular remodeling and may therefore represent useful targets to prevent or treat complications of CKD.

Effects of sevelamer treatment on cardiovascular abnormalities in mice with chronic renal failure.

Elevated serum phosphate and fibroblast growth factor 23 (FGF23) levels are associated with cardiovascular disease (CVD) in patients with chronic renal failure (CRF). The phosphate-binder sevelamer has been shown to decrease both phosphate and FGF23, but limited data indicate that sevelamer improves CRF-related CVD, such as diastolic dysfunction, left ventricular hypertrophy (LVH), and aortic stiffness. To gain additional information, we measured the effects of sevelamer on CVD in a murine model of CRF. Groups of CRF and sham-operated mice received regular chow or 3% sevelamer-HCl in the chow for 14 weeks, starting 6 weeks after the initiation of CRF or sham operation. After the first 8 weeks of sevelamer treatment, CRF mice had decreased serum phosphate levels and an improved aortic systolic expansion rate, pulse-wave velocity, and diastolic function, although LVH remained unchanged. Following an additional 6-week course of sevelamer, LVH had not progressed. The FGF23 serum level was not reduced by sevelamer until after 14 weeks of treatment. In multiple regression analysis, serum phosphate, but not FGF23, was independently correlated with LV diastolic function and mass. Thus, sevelamer first improved aortic stiffness and diastolic dysfunction and secondarily prevented LVH in mice with CRF. The phosphate-lowering, rather than FGF23-lowering, effect appears to be responsible for the observed cardiovascular improvement.

Upregulation of BAD, a pro-apoptotic protein of the BCL2 family, in vascular smooth muscle cells exposed to uremic conditions.

Chronic kidney disease (CKD) has recently emerged as a major risk factor for cardiovascular pathology. CKD patients display accelerated atherosclerotic process, leading to circulatory complications. However, it is currently not clear how uremic conditions accelerate atherosclerosis. Apoptosis is an important homeostatic regulator of vascular smooth cells under pathological conditions. In the present study, we explored the regulation of apoptosis in cells of the vascular wall in the uremic context. We analysed the expression and regulation of the proteins of the BCL2 family that play an essential role in apoptosis. Our results, obtained in mice and primary human smooth muscle cells exposed to two uremic toxins, point to the existence of an alteration in expression and function of one pro-apoptotic member of this family, the protein BAD. We explore the regulation of BAD by uremic toxins and report the sensitization of vascular smooth muscle cells to apoptosis upon BAD induction.

Mechanisms of aortic and cardiac dysfunction in uremic mice with aortic calcification.

BACKGROUND: Chronic renal failure (CRF) is associated with cardiac dysfunction and increased aortic stiffness. The mechanisms involved are not clearly understood. We examined changes over time in cardiac and aortic function in a murine CRF model. METHODS AND RESULTS: Eight-week-old mice were randomly assigned to 1 of 4 groups: wild-type non-CRF, wild-type CRF, apolipoprotein E knockout non-CRF, and apolipoprotein E knockout CRF. Echocardiography was performed and blood samples were taken at baseline and after 6 and 10 weeks of CRF. Vascular reactivity and adhesion molecule expression were studied after 6 and 10 weeks of CRF. Left ventricular hypertrophy, altered left ventricular relaxation, and increased aortic stiffness were observed after 6 weeks of CRF and persisted after 10 weeks. The 4 groups of mice did not significantly differ in terms of arterial blood pressure and aortic structure. The degree of vascular calcification and serum total cholesterol concentration were higher in the CRF groups than in the non-CRF groups. These changes, however, could not explain the cardiac and vascular differences seen in the 2 CRF groups. In contrast, alterations in vascular reactivity, the upregulation of adhesion molecule expression, and CRF status were significantly associated with these changes. CONCLUSIONS: In a mouse model of CRF, left ventricular hypertrophy, cardiac diastolic dysfunction, and increased aortic stiffness were not related to structural changes in the aorta (including aortic calcification) or high serum cholesterol levels. However, cardiac and aortic abnormalities were associated with the extent of subendothelial dysfunction and the severity of CRF.