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Fibroblast growth factor (FGF) 23 and Klotho are two factors associated with several metabolic disorders. Similar to humans, accelerated aging processes characterized by chronic vascular disease, bone demineralization, skin atrophy and emphysema have been recognized in FGF23-null mice and Klotho-deficient mice. The role of these factors in the control of mineral metabolism homeostasis have been shown recently, particularly at the level of parathyroid cells and also in modulating active vitamin D production, two phenomena which are relevant in the presence of chronic kidney disease. In addition, the hormonal affect of circulating FGF23 and Klotho proteins on vascular reactivity, either directly on endothelial cell functions or indirectly by modulating the brain endothelin-1-dependent sympathetic nervous system activity, has contributed to understanding their role in the pathophysiology of hypertension and atherosclerotic vasculopathies. Consequently, very recent clinical investigations seem to confirm the involvement of Klotho in modulating the severity and prognosis of human cardiovascular (CV) disorders and longevity. The present review reports data related to the possible interactive effects of Klotho and FGF23 on the prognosis of renal and CV diseases.

Background Recent studies have suggested that vitamin D and an imbalance in calcium homeostasis may have an impact on the cardiovascular system. The aim of this study was to assess the impact of different concentrations of extracellular Ca(2+) on human umbilical vein cord endothelial cells (HUVEC) by measuring its effect on parameters involved in the pathogenesis of vascular inflammatory responses. Methods HUVEC were grown in the 3.5, 4.5 or 5.8 mg/dL concentration of extracellular Ca(2+) for 2-3 weeks. Expression of adhesion molecules was analysed by flow cytometry. Endothelial nitric oxide synthase (eNOS), receptor of advanced glycation end-product (RAGE) and interleukin-6 (IL-6) mRNA expressions were determined by real-time PCR. eNOS, inhibitor kappa Balpha (IkappaBalpha) and phosphorylated IkappaBalpha protein levels by Western blot, eNOS activity by conversion of [(14)C]-arginine to [(14)C]-citrulline, IL-6 and osteoprotegerin (OPG) secretion by ELISA and DNA-binding activity of nuclear factor kappa B (NFkappaB)-p65 were assayed colorimetrically in nuclear extracts. Results In the presence of low Ca(2+) (3.5 mg/dL), protein expressions and activity of eNOS were diminished, while the protein expressions of intercellular adhesion molecule-1 (ICAM-1), as well as the mRNA expressions of RAGE and IL-6, were elevated. The protein secretions of IL-6 and OPG were also stimulated in low Ca(2+) concentration. At this concentration, the DNA-binding activity of NFkappaB was enhanced, probably due to the decreased level of IkappaBalpha. Conclusions These results suggest that lower extracellular ionized Ca(2+) may play a relevant role in modifying endothelial cells functions.

BACKGROUND Parathyroid hormone (PTH), the major systemic calcium regulating hormone has been implicated in the development of hypertension and the occurrence of uraemic vascular changes. As nitric oxide synthase (NOS) is involved in the production of nitric oxide, and acute PTH effect is characterized by vasodilation, the effect of PTH on the endothelial NOS (eNOS) system was measured in cultured human umbilical cord vein endothelial cells (HUVEC) and the pathways possibly involved were studied. METHODS The presence of the PTH receptor-1 (PTHR1) on the HUVEC membrane was examined by RT-PCR, immunocytochemistry and western blot. HUVEC were stimulated with 10(-12) to 10(-10) mol/l PTH. The eNOS mRNA expression was established by RT-PCR and the eNOS protein levels were determined by western blot. The eNOS activity was measured by the conversion of [(14)C]arginine to [(14)C]citrulline. RESULTS PTHR1 has been found to be expressed in HUVEC and its expression is depressed by increasing concentrations of PTH. PTH induced a significant increase in eNOS mRNA (10(-11) mol/l: 1.87 +/- 0.16, P = 0.012; 10(-10) mol/l: 1.96 +/- 0.28, P = 0.007, fold of control), and protein expression. The eNOS activity was also significantly stimulated (10(-11) mol/l: 1139 +/- 203; 10(-10) mol/l: 1323 +/- 216 vs control: 621 +/- 154 cpm/150 mug protein, P < 0.01). The addition of calphostin C (PKC inhibitor) or Rp-cAMP (PKA inhibitor) reduced the eNOS mRNA, protein expression and activity of PTH-stimulated HUVEC. The combined treatment of calphostin C and Rp-cAMP abolished the eNOS protein expression and activity. CONCLUSION PTH induces an increased activity of the eNOS system; probably both PKA and PKC pathways are involved in this activation. Such data may explain the vasodilation observed after acute treatment with PTH.

Background. Chronic treatment with candesartan cilexetil (C) improves the outcome of rats after 5/6 nephrectomy (Nx). Tetrahydrobiopterin (BH4), an essential cofactor for appropriate endothelial nitric oxide synthase (eNOS) activity, prevents an increase in blood pressure (BP) in Nx rats when given immediately after surgery. In the present study, we evaluated the renoprotective effect of a combined treatment. Methods. Five groups of rats were studied: SHAM (sham-operated rats, n ¼ 12); SNx (untreated 5/6 nephrectomized rats, n ¼ 15); C (SNx rats treated with candesartan cilexetil, 5 mg/kg/day per os, n ¼ 11); C þ BH4 (SNx rats treated with candesartan cilexetil and BH4, 10 mg/kg/day intraperitoneally, n ¼ 11); and BH4 (SNx rats treated with BH4, 10 mg/ kg/day intraperitoneally, n ¼ 11). Treatment began 30 days after surgery, when hypertension and renal insufficiency have developed. This day was considered as day 1 of treatment for statistical comparisons. The study was continued until 50% mortality was achieved in the SNx rats (4 months after surgery). Results. The survival rates were 100% for SHAM, 47% for SNx, 50% for BH4, 64% for C and 80% for C þ BH4 (P < 0.05 vs all). Untreated Nx rats developed hypertension, proteinuria (UP) and severe renal insufficiency. Mortality was associated with a lower renal function and increased urine protein excretion. In C and C þ BH4 rats, systolic blood pressure (SBP) decreased significantly. BH4 alone had a mild nonsignificant effect on SBP. C and C þ BH4 treatments attenuated significantly the increase in proteinuria found in SNx animals. The weight of the remnant kidneys as well as the severity of glomerulosclerosis were significantly lower in the C þ BH4 rats. Conclusion. This study shows that in subnephrectomized rats, addition of BH4 to a treatment with candesartan had an additive renoprotective effect. The mechanism of such action may include a better control of BP associated with a blockade of actions of angiotensin II (Ag II), an improvement in nitric oxide synthesis and a balanced redox.

Background Current blood pressure (BP) classification is based on the recent recommendations of the Joint National Committee on Prevention, Detection, Evaluation and Treatment of High Blood Pressure (JNC-7) and the 2003 European Society of Hypertension–European Society of Cardiology Guidelines for the Management of Arterial Hypertension. The JNC-7 introduced a new concept, prehypertension, and recommended health-promoting lifestyle modifications for these individuals. Obesity is also recognized as a major risk factor for the development of hypertension. We aimed to determine the prevalence of hypertension and obesity in a large cohort of adolescents and to assess whether prehypertension and body mass index (BMI) increase with increasing age. Methods A cross-sectional population-based study was performed using data collected during 1996 to 2002 in an army recruitment examination of 560,588 Israeli individuals 16.5 to 19 years of age. The subjects were divided according to gender and stratified by increasing 6-month intervals into five groups. Prehypertension was defined as BP 120 to 139 / 80 to 89 mm Hg. Overweight was defined as BMI 25 to ≤30 and obesity as BMI >30 kg/m2. Results Mean systolic BP (SBP) and diastolic BP (DBP) were significantly higher in male subjects for all groups. By applying the JNC-7 criteria, 56.8% of male subjects and 35.8% of female subjects would be considered prehypertensive. There was a statistically significant increase in the mean SBP and DBP with age and BMI. Among male subjects 10.9% were overweight and 3.3% were obese; among female subjects, 11.1% were overweight and 3.2% were obese. The BMI did not increase with increasing age. The prevalence of prehypertension was significantly higher in obese subjects. Conclusions Prehypertension is very common among Israeli adolescents. A substantial number of adolescents exhibit a BMI greater than normal. As both of these factors are known to be asssociated with increased cardiovascular risk, early institution of healthful lifestyle changes in a large proportion of this age group is recommended.

Background. The renoprotective effect of ACE inhibition in chronic renal disease is well established but the studies on effects of calcium antagonists on progression of renal disease and on proteinuria have given varying results. Methods. We conducted an open long-term randomized prospective multi-centre study comparing the combination of ramipril (R) and felodipine ER (F) with either drug alone in non-diabetic renal disease. Included were patients with uncontrolled hypertension (diastolic blood pressure (DBP)) P95 mmHg on treatment with a diuretic and a beta-blocker. Fiftyone patients received the combination of R and F, 54 patients R, and 53 patients F. The treatment goal was aD BP-90 mmHg and a similar BP reduction in the three groups. Mean doses at the last visit were 5q5, 10 and 9 mg, respectively, after a mean treatment time of nearly 2 years. The progression of renal impairment was studied by serial measurements of serum creatinine, iohexol clearance, and albuminuria. Results. The reduction in supine systolic (S) BP and DBP expressed as median values were 19.0u14.5, 14.3u15.0 and 13.5u13.3 mmHg in the RqF, R, and F groups, respectively. There was no significant difference between the groups. When correction for the acute drug effect was performed the RqF group had a slower progression rate of the renal disease (loss of glomerular filtration rate (GFR) mluminuyear) compared with the F group (P-0.05) but not to the R group (P)0.20). There was a rise in albuminuria after 2 years in the F group (P-0.05), but no significant change was found in the other groups. Conclusions. In patients with non-diabetic renal disease the combination of an ACE inhibitor and a calcium antagonist in reduced doses used in addition to baseline therapy with beta-blockers and diuretics, tended to cause a better BP reduction as each drug per se. The RqF treatment also caused a slower progression of the renal disease compared with F alone. The combination treatment seems to afford better BP control and appears to be a favourable therapeutic option in patients with renal disease and hypertension.

Background. Pregnant rats with adriamycin nephropathy (ADRP rats) develop hypertension and have an increased vascular reactivity to noradrenaline in the isolated mesenteric bed in vitro. We have shown previ- Introduction ously that the administration of daltroban, a specific thromboxane receptor antagonist, prevented hyperten- Hypertension in pregnancy is associated with increased sion in ADRP rats. vascular reactivity, compared with the decreased vascuMethods. We measured the eVect of daltroban (10’5 lar resistance observed during normal pregnancy [1]. mol/l ) on the vasoconstrictory response to noradrena- The clinical picture of pre-eclampsia displays many line (1‐10 mmol/l ) in the isolated mesenteric bed of functional features of endothelial disease [2]. Recent ADRP rats at the end of pregnancy, as compared with studies have shown that blood pressure increased in normal pregnant and adriamycin-treated virgin rats. pregnant rats with early adriamycin nephropathy In further experiments, we measured the changes of (ADRP rats). This was accompanied by a worsening flow induced by increasing concentrations of the of proteinuria and a relative decrease of glomerular thromboxane analogue, U46619 (10’7‐10’6 mol/ l) . filtration rate (GFR) [3]. The development of hyperFinally, changes of flow were assessed in arteries tension in pregnancy, in contrast with the decrease of maximally constricted with U46619 (10’6 mol/ l), blood pressure occurring during pregnancy in other during perfusion in the presence of increasing concen- hypertensive rat models [4‐8], rendered this a specific trations of daltroban (10’7‐10’5 mol/ l). model and prompted us to look for possible mechanResults. Daltroban diminished the response to nor- isms of deranged vascular behaviour. In previous adrenaline in all groups, shifting the concentration- works, it has been demonstrated that hypertension is eVect curve to the right. However, at maximal concen- completely prevented by treatment with a thromboxane trations of noradrenaline, daltroban was ineVective in receptor antagonist [9]. Subsequently it has been all rats, except in ADRP animals. The vasoconstrictory shown that in ADRP rats, the vascular reactivity of response to U46619 was significantly reduced in all isolated perfused mesenteric beds to noradrenaline was pregnant rats, both normal and adriamycin-treated. not decreased as expected during normal pregnancy Daltroban progressively released the vasoconstriction [10]. In the present study, we have evaluated the induced by U46619 in all groups. However, this vaso- possible role played by thromboxane in the altered dilator response was attenuated in the adriamycin- response of the isolated mesenteric vasculature of treated rats, the slopes of their curves being smaller ADRP rats to the vasoconstrictive action of than those of the respective untreated groups noradrenaline. (0.038±0.006 in virgin rats vs 0.063±0.011 in controls, P

The rat remnant kidney, a popular model of experimental renal failure, has also been used to assess the histological bone changes associated with reduction in renal mass. The suitability of this model has been challenged, especially with regard to the standardization of animals from different age groups and various degrees of renal failure. The present study was undertaken: (1) to better evaluate the histomorphometric changes associated with reduction in renal mass ; (2) to assess these changes over a longer follow-up period as compared to those of matched intact animals ; (3) to define the response to pharmacological doses of calcitriol. Male rats were evaluated 4 and 8 weeks after induction of renal failure (5/6 nephrectomy) and compared with age-matched control rats with intact kidneys. Plasma biochemistry, creatinine clearance and parathyroid hormone (PTH ) were obtained from normal rats and from the rats with chronic renal insufficiency. Histomorphometric studies were performed in all animals on the right tibial bone. Induction of renal failure of 4 weeks' duration (short-term study, age 16 weeks) was associated with increased PTH and bone resorption, but suppressed bone formation. At long-term follow-up (rats aged 20 weeks), the suppression in bone formation was even more prominent. These changes were assessed considering normal bone histomorphometry during the process of growing. Calcitriol administration was associated with a significant suppression of bone resorption and a spectacular increase in all osteoid parameters. The bone formation rate, however, remained in the same low range as that in untreated animals. This model of renal osteodystrophy is similar to human secondary hyperparathyroidism with regard to the enhanced bone resorption, but differs by the marked decrease in bone formation. There was a remarkable similarity between the response of rats with renal failure and that of patients with severe secondary hyperparathyroidism to large doses of calcitriol, leading in both to adynamic bone disease. Keeping in mind the species differences, the bone changes developing in the uraemic rats may be useful in understanding certain aspects of human renal osteodystrophy.