Rika Jimbo

Fibroblast growth factor 23 accelerates phosphate-induced vascular calcification in the absence of Klotho deficiency

Fibroblast growth factor 23 (FGF23) is a phosphate-regulating hormone that acts primarily on the kidney and parathyroid. With declining kidney function there is an increase in circulating FGF23 levels, which is associated with vascular calcification and mortality in chronic kidney disease. Whether FGF23 exerts direct effects on vasculature is unclear. We evaluated the expression of Klotho and FGF receptors in rat aortic rings and rat aorta vascular smooth muscle cells maintained in culture by reverse transcription-PCR, western blotting, and immunostaining. Signaling pathways underlying FGF23 effects were assessed by western blotting, and effects of FGF23 on osteogenic markers and phosphate transporters were assessed by real-time reverse transcription-PCR. We detected Klotho and FGFR1 in total aorta but not in vascular smooth muscle cells. FGF23 augmented phosphate-induced vascular calcification in the aortic rings from uremic rats and dose dependently increased ERK1/2 phosphorylation in Klotho-overexpressing but not naive vascular smooth muscle cells. FGF23-induced ERK1/2 phosphorylation was inhibited by SU5402 (FGFR1 inhibitor) and U0126 (MEK inhibitor). FGF23 enhanced phosphate-induced calcification in Klotho-overexpressing vascular smooth muscle cells and increased osteoblastic marker expression, which was inhibited by U0126. In contrast, phosphate transporter expression was not affected by phosphate or FGF23. Thus, FGF23 enhances phosphate-induced vascular calcification by promoting osteoblastic differentiation involving the ERK1/2 pathway.

Cardiovascular Risk Factors and Chronic Kidney Disease—FGF23: A Key Molecule in the Cardiovascular Disease

Patients with chronic kidney disease (CKD) are at increased risk of mortality, mainly from cardiovascular disease. Moreover, abnormal mineral and bone metabolism, the so-called CKD-mineral and bone disorder (MBD), occurs from early stages of CKD. This CKD-MBD presents a strong cardiovascular risk for CKD patients. Discovery of fibroblast growth factor 23 (FGF23) has altered our understanding of CKD-MBD and has revealed more complex cross-talk and endocrine feedback loops between the kidney, parathyroid gland, intestines, and bone. During the past decade, reports of clinical studies have described the association between FGF23 and cardiovascular risks, left ventricular hypertrophy, and vascular calcification. Recent translational reports have described the existence of FGF23-Klotho axis in the vasculature and the causative effect of FGF23 on cardiovascular disease. These findings suggest FGF23 as a promising target for novel therapeutic approaches to improve clinical outcomes of CKD patients.

Oxidative Stress in Multiple Organ Damage in Hypertension, Diabetes and CKD, Mechanisms and New Therapeutic Possibilities

Hypertension, diabetes, hypercholesterolemia and chronic kidney disease (CKD) lead to cardiovascular (CV) events and cardiovascular death consists of main cause in mortality of those diseases. Understanding of pathophysiology that links them and CV events has been vigorously studied and several factors are believed to play roles such as NO, reninangiotensin system, and oxidative stress. It has been shown that those factors affect endothelial function and consequently organ circulation as well as function and viability of cells and organs. Despite overwhelming evidences in the consequences of experimental models of ROS-induced organ damage, large-scale clinical trials of former antioxidant therapies, such as vitamin C, vitamin E or -carotene, could not demonstrate satisfactory benefit to patients and they seemed to be harmful in some cases (Hennekens et al., 1996; Omenn et al., 1996; Virtamo et al., 1998; Hercberg et al., 1999; Lee et al., 1999; Yusuf et al., 2000; de Gaetano, 2001; Heart, 2002; Vivekananthan et al., 2003; Hercberg et al., 2004; KrisEtherton et al., 2004; Lonn et al., 2005). Several studies concluded that β-carotene supplementation increased the relative risk of death in patients with some types of cancer and had no benefit on patients with cardiovascular disease. Another study said vitamin E increased hemorrhagic stroke. Even antioxidant cocktails increased in all-cause motality (Omenn et al., 1996; Rosen et al., 2001a). So far, supplementation with vitamins C and E, either alone or in combination with each other or with other antioxidant vitamins, does not appear to be efficacious for the treatment of cardiovascular disease (Lonn et al., 2005). We investigated role of oxidative stress in consequences of multiple organ damages in mouse and possible new therapeutic agent.

Phosphorus and Mortality: Do We Have the Panacea?

Epidemiological studies show that serum phosphate concentration is an independent risk factor for cardiovascular morbidity and death in CKD patients. The effects of phosphate on the cardiovascular system have been extensively studied, and concurrent perturbations in the FGF23—Klotho axis have been shown to play a pivotal role in this process. To reduce cardiovascular complications and death, pharmacological as well as dietary interventions to reduce serum phosphate levels are key. On a population basis, an overall restriction of dietary intake of phosphorus is likely to yield promising effects on cardiovascular outcomes, in part by altering FGF 23 and Klotho dynamics.

Renal Denervation Improves Cardiac Diastolic Dysfunction by Restoring Serca2a Transcription in Uninephrectomized Rats

Background: The mortality and morbidity of heart failure with preserved ejection fraction has increased. Sarcoplasmic reticulum Ca2+-ATPase type 2a (SERCA2a) regulates cardiac functions, and a reduction in SERCA2a expression has been documented in left ventricular (LV) diastolic dysfunction. By contrast, SERCA2a overexpression improves LV diastolic dysfunction. Thus, transcriptional regulation of SERCA2a may be a new therapeutic target. The aim of this study was to determine whether renal denervation, a treatment for resistant hypertension, is a regulator of SERCA2a transcription in vivo. Methods: Uninephrectomy and 6-week salt loading in three-week-old male Sprague-Dawley rats were used to devise a cardiac diastolic dysfunction model, and mechanical renal denervation was performed. The expression of SERCA2a and related molecules was evaluated with quantitative polymerase chain reaction and western blot analyses. The maximal positive LV pressure development (+dP/dtmax) and time constant at the isovolumic relaxation phase (Tau) were determined with cardiac catheters. Results: Uninephrectomy combined with a high-salt diet significantly reduced the messenger RNA expression and protein abundance of SERCA2a, which were restored by renal denervation. In accordance with changes in SERCA2a transcription, uninephrectomy and the high-salt diet decreased LV diastolic function, which was evaluated by Tau and restored by renal denervation. LV systolic function, measured with +dp/dtmax, was preserved. Renal denervation did not lower blood pressure, urinary protein levels, cardiac hypertrophy, or fibrosis. Conclusions: We found that renal denervation is a regulator of SERCA2a transcription in vivo. Our data may provide new therapeutic insights into LV diastolic dysfunction and warrant further study.

Ps 10-15 Torasemide Dual Blockade Of Pge Synthesis And Sodium Channel In Renal Failure Model

Objective: Torasemide is a long-acting loop diuretic. Previous study showed that torasemide is associated with low mortality in patients with chronic heart failure (CHF). But it is not known if torasemide is superior in protecting kidney to furosemide. Renal blood flow is precisely controlled to maintain glomerular filtration and fluid and electrolyte homeostasis. Eicosanoids play a critical role in the control of renal hemodynamic as well as inflammations and sodium balance. There are possibilities that torasemide may modulate eicosanoid pathway and be protective in renal function. Design and Method: Three-week-old uninephrectomized (UNx) Sprague-Dawley (SD) rats were divided into the following groups: 1) Normal salt diet (0.5% NaCl) as a control; 2) HS (8.0% NaCl) diet alone; 3) HS diet plus treatment with the Torasemide (3mg /(kg day), T3); 4) HS diet plus treatment with the Torasemide (5 mg/(kg day), T5); 5) HS diet plus treatment with the Furosemide (30 mg/(kg day), F30); 6) HS diet plus treatment with the Furosemide (100 mg/(kg day), F100); Rats were treated for 4 weeks, body weight, blood pressure (BP), cardiac and renal function is monitored. Results: In current study I used young age of SD rat with high salt loading and uninephrectomized, development of salt-sensitivity hypertension model, after treatment with torasemide decrease body weight, HW/BW, blood pressure (160 mmHg to 120 mmHg), urine protein (125 mg/day to 25 mg/day) level also reverse cardiac left ventricular diastolic function and kidney function. Importantly torasemide could reduce Urinary PGE2 execration level. Conclusions: For the first time we showed that Torasemide have strong protect effect on kidney and cardiac function than Furosemide in salt-sensitive hypertension and CKD model.

588 STRICT FLUID MANAGEMENT PREVENTED LEFT VENTRICULAR HYPERTROPHY IN LONG-TERM PERITONEAL DIALYSIS PATIENTS

Backgrounds: It is still unclear whether strict salt restriction and blood pressure control could prevent left ventricular hypertrophy (LVH) in peritoneal dialysis (PD) patients with declining residual renal function (RRF). Methods: 45 patients who started PD in our hospital between Jan 2005 and July 2010 were investigated retrospectively. Clinical data of these patients including left ventricular mass index (LVMI) evaluated by echocardiographywere studied from the baseline (after 6 months of beginning PD)and thereafter every 6 months. Results: The patients included 32 males and 13 females (age 58.2 ± 11.6 years, observational period 33.7 ± 16.7 months). At baseline, systemic blood pressure (SBP) was 127.6 ± 18.5 mmHg, prevalence of LVH was 46.6% and LVMI was 115.1 ± 32.5 g/m2. We instructed salt restriction less than 6 g/day in daily clinical practice. At the end of follow-up, RRF declined significantly; weekly renal Kt/V from 1.16 ± 0.49 to 0.58 ± 0.54 (P < 0.0001). However, SBP (133.6 ± 20.3 mmHg, P = 0.14) and LVMI (119.7 ± 30.5 g/m2, P = 0.48) were almost unchanged. Stepwise multiple regression analysis showed that SBP (standard &bgr; = 0.607, P < 0.0001), CTR (standard &bgr; = 0.447, P =0.0002) and intact-PTH (standard &bgr; = 0.240, P = 0.02) were independently associated with LVMI (r2 = 0.616). Conclusions: Our report demonstrates for the first time that we could prevent the development of LVH in PD patients with declining RRF, as long as their fluid and blood pressure were kept under good control.

952 FIBROBLAST GROWTH FACTOR 23 ACCELERATES PHOSPHATE-INDUCED CALCIFICATION OF VASCULAR SMOOTH MUSCLE CELLS

Background: Fibroblast growth factor 23 (FGF23) is a phosphate (P) -regulating hormone. Clinical studies suggest that increased level of FGF23 is associated with vascular calcification among chronic kidney disease patients but its direct effect on vasculature is unknown. Methods: Vascular smooth muscle cells (VSMCs) were extracted from Sprague-Dawley rats. The protein expressions of Klotho, co-receptor essential for FGF23 signalling, in VSMCs and the aorta were analyzed by western blot and immunohistochemistry. ERK 1/2 phosphorylation in normal VSMCs or Klotho-overexpressed VSMCs (Klotho-VSMCs) by recombinant hFGF23 was analyzed by western blot. Calcification of Klotho-VSMCs was induced by high-P medium in the presence or absence of FGF23. The calcium depositions were visualized by calcification staining and quantified by methylxylenol blue method. The mRNA expressions of osteoblastic transcription factors and sodium dependent P transporters were evaluated by real-time RT-PCR. Results: Klotho protein was detected in the medial layer of the aorta but not in the VSMCs. ERK 1/2 phosphorylation by FGF23 was increased in Klotho-VSMCs in a dose dependent manner and inhibited by FGFR1 inhibitor or MEK inhibitor U0126. FGF23 accelerated P-induced calcification of Klotho-VSMCs in a dose dependent manner. P induced Msx2 and osterix expressions were enhanced by FGF23 and U0126 inhibited the additive effect of FGF23 on the expression of these osteoblastic markers. The expressions of Pit1 and Pit 2 were not affected by either increased P level or FGF23. Conclusions: FGF23 enhances P-induced calcification in Klotho-VSMCs by promoting osteoblastic differentiation and its effect might be mediated by the ERK 1/2 pathway.

Adrenomedullin and Oxidative Stress in Vascular Damage and Metabolic Disease

We have studied the pleiotropic effects of adrenomedullin with knockout mice and found that it is a potent intrinsic antioxidant. Oxidative stress is related to cardiovascular as well as metabolic diseases. In this review, the coronary, pulmonary, and cerebrovascular damage that was observed in adrenomedullin-knockout mice is reviewed. An angiotensin II-loaded model, a hypoxia model, and a vascular occlusion model were applied to investigate vascular damage, and the results clarified the role of oxidative stress and the therapeutic potency of adrenomedullin in vascular protection. From in vivo and in vitro studies, adrenomedullin is known to antagonize oxidative stress by both inhibiting nicotinamide adenine dinucleotide phosphate oxidase and promoting the degradation of oxidative stress. In addition, the role of adrenomedullin in insulin resistance and obesity is discussed based upon the results of both clinical studies and basic research studies that used aged adrenomedullin-knockout mice. The pleiotropic effects of adrenomedullin suggest a number of new therapeutic targets in a variety of diseases.