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Inverted duplications deletions are rare, complex, and nonrecurrent chromosomal rearrangements associated with a variable phenotype. In this case report, we described the phenotype and genotype of a 14-week-old male fetus, who was aborted after discovery of multiple anomalies (septal cystic hygroma, open abdominal wall, and a nonidentifiable lower limb). At autopsy, fluorescence in situ hybridization and array comparative genomic hybridization identified an inverted duplication with terminal deletion of 4p [46,XY,der(4)del(p16.3)dup(4)(p15.2p16.3)]. Only five genotypically similar cases have been reported, and we hope our case contribution will add meaningful to the body of knowledge.

Several patients with chromosomal deletions including ZFHX4 gene have been described, whereas point mutations are very rare. This gene encodes for a transcription factor involved in the development of several embryonal processes, including brain differentiation. Patients with 8q21.11 deletions usually show intellectual disability, short stature, peculiar facial features, and severe eye abnormalities. We describe a female patient with mild intellectual disability, autism spectrum disorder, strabismus, ptosis, low-set and prominent ears, high-arched palate, microretrognathia. Clinical Exome Sequencing revealed the presence of a de novo heterozygous variant in ZFHX4. Therefore, we further investigate the different phenotypes of ZFHX4 mutations and 8q21.11 deletions.

BACKGROUND: Microgravity induces redistribution of body fluids and reductions in muscle and bone mass. These effects correlate with changes in lab test results, including urea and bone minerals. Difficulties with collecting blood and urine during space missions limit the available data. This pilot study investigated metabolic changes during a space mission using untimed spot samples of urine and saliva. Untimed spot urine was used for urinalysis with data normalization per creatinine concentration. Saliva was proven useful as an index of serum urea and phosphorus.METHODS: Two astronauts collected urine and saliva samples 75 ± 5 d before launch (baseline) and 3-5 times during a 6-mo space mission. Samples were collected 3 h after morning breakfast. Urine was collected using a standard NASA device. Saliva was collected using a Salivette™ synthetic swab. Samples were kept frozen using automated biochemistry until lab work-up. Anthropometric data were collected at baseline and after the mission.RESULTS: For astronauts 1 and 2, respectively, total bone mineral density decreased (-1.4% and -0.9%). In-flight changes were as follows: transiently decreased urine urea/creatinine ratio (-32% and -24%), transiently decreased urine phosphorus/creatinine ratio (-52% and -30%), increased urine calcium/creatinine ratio (up to +116% and +27%), and transient increases in saliva urea (up to +48% and +195%) and phosphorus (up to +29% and +46%). The astronaut with greater changes in urine minerals had greater reduction in bone mineral density.DISCUSSION: The results support the hypothesis that untimed samples of urine and saliva are useful for investigation of metabolic changes during space missions. Changes in urine and saliva minerals suggested down-regulation of parathyroid gland activity during the space mission.Bilancio G, Cavallo P, Lombardi C, Guarino E, Cozza V, Giordano F, Cirillo M. Urea and minerals monitoring in space missions by spot samples of saliva and urine. Aerosp Med Hum Perform. 2019; 90(1):43-47.

Background Lab tests on saliva could be useful because of low invasivity. Previous reports indicated that creatinine, uric acid, and potassium are measurable in saliva. For these analytes the study investigated methodology of saliva tests and correlations between plasma and saliva levels. Methods The study enrolled 15 healthy volunteers for methodological analyses and 42 nephropathic patients for plasma-saliva correlations (35 non-dialysis and 7 dialysis). Saliva was collected by synthetic swap right after venipuncture for blood withdrawal. Blood and saliva, unless otherwise indicated, were collected early in the morning after overnight fast and lab tests were performed in fresh samples by automated biochemistry (standard). Methodological analyses included blind duplicates, different collection mouth sites, day-to-day variability, different collection times, and freezing-thawing effects. Analyses on plasma-saliva correlations included post-dialysis changes. Results For saliva lab tests of all analytes, blind duplicates, samples from different mouth sites or of different days were not significantly different but were significantly correlated (differences ≤14.4%; R ≥ 0.620, P ≤ 0.01). For all analytes, mid-morning saliva had lower levels than but correlated with standard saliva (differences ≥15.8%; R ≥ 0.728, P ≤ 0.01). Frozen-thawed saliva had lower levels than fresh saliva for uric acid only (− 17.2%, P

Background. Research data are limited about indices of osmotic equilibrium and of kidney concentrating activity. This study investigated correlates and prognostic power of these indices in a sample of the general population. Methods. Urine osmolality (U-osm), plasma osmolality (P-osm), plasma creatinine, and other variables were measured by Gubbio Study for the 1988-92 exam (baseline). Plasma creatinine and other variables were re-measured in 2001-07 (follow-up). Kidney concentrating activity was assessed as U-osm/P-osm ratio and kidney function as estimated glomerular filtration rate (eGFR). Results. Baseline data were complete in 4220 adults of whom 852 dead before follow-up and 2795 participated in follow-up. At baseline, there were the following independent cross-sectional associations: female sex and higher urine flow with lower values of U-osm, P-osm, and U-osm/P-osm ratio (P

OBJECTIVE: This population-based study investigated low protein intake, mortality, and kidney function decline. DESIGN: Observational longitudinal cohort study. SUBJECTS: Target cohort consisted of 4,679 adults participating in 1988-1992 and 2001-2007 examinations of the Gubbio Study (baseline and follow-up). Data collection included overnight urine urea nitrogen (UUN) and other variables at baseline, serum creatinine at baseline and follow-up, and mortality from baseline to follow-up. Three hundred seventy-two persons were excluded for missing data. UUN in the lowest 20% of the distribution was defined as low and used as index of low protein intake. Estimated glomerular filtration rate (eGFR, mL/minute × 1.73 m2) was used as kidney function index. INTERVENTION: None (observational study). MAIN OUTCOME MEASURE: Mortality and eGFR decline are the main outcome measures, and eGFR decline was defined as eGFR change from baseline to follow-up ≤ mean-1 standard deviation (Z-score ≤ -1). RESULTS: Eight hundred seventy-one deaths occurred over 15.9 ± 4.0 years of observation (417 from cardiovascular disease and 276 from neoplastic disease). Low UUN associated with mortality (hazard ratio, HR = 1.31, 95% confidence interval, CI = 1.12/1.53) due to association with mortality from neoplastic disease (HR = 1.33, 95% CI = 1.02/1.76). Mortality-corrected follow-up response rate was 79.9% (n = 2845). Baseline to follow-up eGFR change was -9.9 ± 10.1, and eGFR decline was found in 454 examinees. Low UUN associated with eGFR decline only in subgroup with baseline eGFR

BACKGROUND: Kidney function measured as estimated glomerular filtration rate (eGFR) is a risk factor for mortality and severe diseases. Protein intake up-regulates kidney function. The dose-response curve of eGFR over protein intake is unknown. Urinary urea nitrogen is an objective index of protein intake. METHODS: The study cross-sectionally analysed the relation between overnight urinary urea nitrogen ((on)U-ureaN) and eGFR with and without control for other variables in 4106 adults of the Gubbio population. Analyses were done for serum creatinine (S-cr) also to investigate the independency of results from eGFR calculation. RESULTS: Higher (on)U-ureaN associated with higher eGFR, and lower S-cr independently of sex and age (simple and partial correlation coefficients >0.100, P < 0.001). Analyses by (on)U-ureaN decile indicated sigmoid curves of eGFR and S-cr over (on)U-ureaN with trend to flatness in the lowest 20% and the highest 20% of (on)U-ureaN (10.12 mg/h, respectively). Multi-variable spline regression indicated that the relation of eGFR over (on)U-ureaN was non-significant for (on)U-ureaN 10.12 mg/h (coefficient = +0.05, 95% CI = -0.06/ +0.16, P = 0.394). eGFR differed by ≈8 mL/min × 1.73 m(2) between the lowest and highest 20% of (on)U-ureaN distribution. CONCLUSIONS: Higher protein intake relates to higher eGFR. The relation is sigmoid with eGFR up-regulation for (on)U-ureaN >5.19 mg/h, a threshold approximately corresponding to the recommended daily allowance for protein intake (0.8 g/day per kg of ideal weight).

BACKGROUND: Phosphorus and urea are measurable in saliva. Measurements of saliva phosphorus (S-Pho) and saliva urea (S-Urea) could be useful because of low invasivity. Data are limited to saliva tests methodology and to correlations between plasma and saliva compositions. S-Pho and S-Urea were investigated focusing on blind duplicates, differences between collection sites, differences between collection times, freezing-thawing effects, and plasma-saliva correlations. METHODS: Tests were performed using fresh saliva collected by synthetic swap early morning after overnight fast (standard). Methodology was investigated in fifteen healthy volunteers. Plasma-saliva correlations were investigated in thirty nephropathic outpatients. RESULTS: S-Pho and S-Urea in all measurements ranged above detection limits (0.3 mmol/L). In healthy volunteers, S-Pho and S-Urea were similar in duplicates (results for S-Pho and S-Urea: % difference between samples ≤ 4.85%; R between samples ≥ .976, P < .001), in samples from different mouth sites (≤4.24%; R ≥ .887, P < .001), and in samples of different days (≤5.61%; R ≥ .606, P < .01) but, compared to standard, were substantially lower in after-breakfast samples (-28.0% and -21.3%; R ≥ .786, P < .001) and slightly lower in frozen-thawed samples (-12.4% and -5.92%; R ≥ .742, P < .001). In nephropathic patients, S-Pho was higher than but correlated with plasma phosphorus (saliva/plasma ratio 4.80; R = .686, P < .001), whereas S-Urea and plasma urea were similar and correlated with each other (saliva/plasma ratio 0.96; R = .944, P < .001). Post-dialysis changes in S-Pho and S-Urea paralleled post-dialysis changes in plasma phosphorus and urea. CONCLUSION: S-Pho and S-Urea reflect plasma phosphorus and plasma urea. Early morning fasting fresh samples are advisable because collection time and freezing-thawing affect saliva tests.

Short-term effects of chlorthalidone are unknown in low kidney function. The effects of 8-week treatment with 25-mg chlorthalidone on the top of ongoing treatment were compared between control hypertensives and low kidney function hypertensives as assessed by estimated glomerular filtration rate 2 . Screening period consisted of 2 visits for patient selection and pretreatment laboratory evaluations (baseline). Inclusion criteria were uncontrolled hypertension on nondiuretic antihypertensive treatment. Exclusion criteria were chlorthalidone contraindications, refused consent, treatment with >3 antihypertensive drugs, severe hypertension, severe comorbidities, unreliable estimated glomerular filtration rate. Treatment period consisted of 5 visits (weeks 1, 2, 4, 6, and 8). Post-treatment laboratory evaluations were performed 3 to 4 days before week-8 visit. The 2 groups differed for baseline estimated glomerular filtration rate (low kidney function and control: n=60 and 60; mean, 39 and 76; range, 15–59 and 60–104) but not for sex, age, and baseline blood pressure. Week-8 blood pressure changes were a decrease in both groups (low kidney function and control: systolic pressure, −20 and −23; 95% confidence interval, −22/−18 and −26/−19; diastolic pressure, −9 and −10, −11/−7, and −13/−8) without significant between-group differences. Incidence of adverse events was similar in the 2 groups (15.0% and 16.7%). Baseline estimated glomerular filtration rate did not predict blood pressure changes and adverse events in either groups ( P >0.6). In both groups, post-treatment changes were a decrease for estimated glomerular filtration rate and serum potassium, an increase for serum uric acid ( P

Opinions are conflicting about the epidemiology of vitamin D deficiency. This population-based study investigated cross-sectionally the associations of 25-hydroxyvitamin D (calcidiol) and 1,25-dihydroxyvitamin D (calcitriol) with indices of mineral homeostasis. Study cohort consisted of 979 persons of the Moli-Sani study, both sexes, ages &ge, 35 years. Data collection included serum calcidiol by different assays, serum calcitriol, serum parathyroid hormone, serum and urine calcium, and phosphorus. Prevalence of mild-to-moderate calcidiol deficiency (10&ndash, 19 ng/mL) was 36.4% and did not associate with hypocalcemia or hyperparathyroidism. Prevalence of severe calcidiol deficiency (&lt, 10 ng/mL) was 16.8% and associated with hyperparathyroidism only (odds ratio = 8.81, 95% confidence interval = 2.4/32.9). Prevalence of calcitriol deficiency (&lt, 18 pg/mL) was 3.1% and associated with hypocalcemia (29.1, 7.4/114.5) but not hyperparathyroidism. In ANOVA along concentration strata, lower calcidiol associated with higher parathyroid hormone only (p &lt, 0.001). Lower calcitriol associated with lower serum and urine calcium (p &lt, 0.001) but not with parathyroid hormone. Calcidiol findings were consistent with different calcidiol assays. In the population, mild-to-moderate calcidiol deficiency did not associate with abnormal mineral homeostasis. Severe calcidiol deficiency and calcitriol deficiency associated with different disorders: lower calcidiol associated with hyperparathyroidism whereas lower calcitriol associated with hypocalcemia and low urine calcium.

Partial trisomy 16 is rare and most of the reported cases are secondary to chromosome rearrangements resulting in concurrent monosomies or trisomies of a second chromosome. Only a few patients survive the neonatal period and the duplication of the long arm seems to be mainly responsible for the prenatal lethality of the full trisomy 16. The reported patients with a partial 16q trisomy have a wide spectrum of congenital anomalies that include dysmorphic features, central nervous system malformations, failure to thrive, and club feet. The patients with duplications of proximal 16q frequently have short stature, developmental delay, speech delay, learning difficulties, and mild to severe behavioral problems. Here we describe a patient with an inverted de novo tandem duplication of 16q with breakpoints evaluated in detail by molecular-cytogenetic techniques. Main clinical features include postural, motor and speech delay with severe learning difficulties and behavioral problems, obesity, microcephaly, and mild dysmorphic features. In the report we attempt to classify the few reported patients with pure partial duplications of 16q in more narrow and homogeneous groups: proximal, proximal-intermediate, intermediate, and intermediate-distal duplications. Moreover, we emphasize the importance of proper cytogenetic investigation and complete molecular cytogenetic refinement in all cases with a suspected chromosomal anomaly.

Hydrogen sulfide, H2S, is the third endogenous gas with cardiovascular properties, after nitric oxide and carbon monoxide. H2S is a potent vasorelaxant, and its deficiency is implicated in the pathogenesis of hypertension and atherosclerosis. Cystathionine beta-synthase, cystathionine gamma- lyase, and 3-mercaptopyruvate sulfurtransferase catalyze H2S formation. Chronic kidney disease is characterized by high prevalence of hyperhomocysteinemia, hypertension, and high cardiovascular mortality, especially in hemodialysis patients. H2S levels are decreased in hemodialysis patients through transcriptional deregulation of genes encoding for the H2S-producing enzymes. Potential implications relate to the pathogenesis of the manifestations of the uremic syndrome, such as hypertension and atherosclerosis. (C) 2010 by the National Kidney Foundation, Inc. All rights reserved.

The routine prenatal diagnosis includes a standard cytogenetic study and a careful evaluation of the fetus by one or more ultrasonographic scans. Usually there is a full concordance between cytogenetic and ultrasonographic fetal sex determination, but occasionally a discordant result may be observed (1/2500 pregnancies). In these cases, besides a possible mistake and an exchange between amniotic fluid samples, a disorder of sex determination should be considered. Sex reversal can be divided into 46,XY female sex reversal syndrome and 46,XX male sex reversal syndrome. The XX male syndrome (OMIM 278850) is a rare condition affecting 1 in 20 000 newborn males. Molecular analysis of sex-reversed patients led to the discovery of the SRY gene (sex-determining region on Y), a gene playing a crucial role in the male differentiation of the bipotential gonad (Sinclair et al., 1990).

Hyperhomocysteinemia is an independent cardiovascular risk factor, according to most observational studies and to studies using the Mendelian randomization approach, utilizing the common polymorphism C677T of methylene tetrahydrofolate reductase. In contrast, the most recent secondary preventive intervention studies, in the general population and in chronic kidney disease (CKD) and uremia, which are all negative (with the possible notable exception of stroke), point to other directions. However, all trials use folic acid in various dosages as a means to reduce homocysteine levels, with the addition of vitamins B6 and B12. It is possible that folic acid has negative effects, which offset the benefits; alternatively, homocysteine could be an innocent by-stander, or a surrogate of the real culprit. The latter possibility leads us to the search for potential candidates. First, the accumulation of homocysteine in blood leads to an intracellular increase of S-adenosylhomocysteine (AdoHcy), a powerful competitive methyltransferase inhibitor, which by itself is considered a predictor of cardiovascular events. DNA methyltransferases are among the principal targets of hyperhomocysteinemia, as studies in several cell culture and animal models, as well as in humans, show. In CKD and in uremia, hyperhomocysteinemia and high intracellular AdoHcy are present and are associated with abnormal allelic expression of genes regulated through methylation, such as imprinted genes, and pseudoautosomal genes, thus pointing to epigenetic dysregulation. These alterations are susceptible to reversal upon homocysteine-lowering therapy obtained through folate administration. Second, it has to be kept in mind that homocysteine is mainly protein-bound, and its effects could be linked therefore to protein homocysteinylation. In this respect, increased protein homocysteinylation has been found in uremia, leading to alterations in protein function. © 2009 Wiley Periodicals, Inc.

Plasma proteins in hemodialysis patients display a significant increase in deamidated/isomerized Asx (asparagine and aspartic acid) content, a marker of protein fatigue damage. This has been linked to the toxic effects of hyperhomocysteinemia in uremic erythrocytes; however, treatment aimed at abating homocysteine levels did not lead to significant reductions in plasma protein damage. The hypothesis that lack of reduction in protein damage could be due to protein increased intrinsic instability, as result of interference with the uremic milieu rather than to hyperhomocysteinemia, was put forward. The deamidated/isomerized Asx content of normal plasma incubated with several uremic toxins for 24 h, 72 h, and 7 d was measured, identifying a group of toxins that were able to elicit this kind of damage. Uremic toxins were also incubated with purified human albumin, and dose-response experiments with the two most toxic agents in terms of protein damage (guanidine and guanidinopropionic acid) were carried out. The effect of the hemodialysis procedure on protein damage was evaluated. For investigating also the consequences of these alterations, human albumin was treated in vitro to produce an increase in deamidated/isomerized Asx residues, and the effects of albumin deamidation on protein binding were evaluated. Among the uremic toxins that are able to elicit protein damage, guanidine produced a dose-dependent increase in protein damage. No difference was found after a hemodialysis session. Deamidated albumin shows normal binding capacity to warfarin, salicylic acid, or diazepam but reduced binding to homocysteine. In conclusion, uremic toxins, especially guanidine, display an ability to induce significant protein damage, which can in turn have functional consequences. Plasma proteins in hemodialysis patients display a significant increase in deamidated/isomerized Asx (asparagine and aspartic acid) content, a marker of protein fatigue damage. This has been linked to the toxic effects of hyperhomocysteinemia in uremic erythrocytes; however, treatment aimed at abating homocysteine levels did not lead to significant reductions in plasma protein damage. The hypothesis that lack of reduction in protein damage could be due to protein increased intrinsic instability, as result of interference with the uremic milieu rather than to hyperhomocysteinemia, was put forward. The deamidated/isomerized Asx content of normal plasma incubated with several uremic toxins for 24 h, 72 h, and 7 d was measured, identifying a group of toxins that were able to elicit this kind of damage. Uremic toxins were also incubated with purified human albumin, and dose-response experiments with the two most toxic agents in terms of protein damage (guanidine and guanidinopropionic acid) were carried out. The effect of the hemodialysis procedure on protein damage was evaluated. For investigating also the consequences of these alterations, human albumin was treated in vitro to produce an increase in deamidated/isomerized Asx residues, and the effects of albumin deamidation on protein binding were evaluated. Among the uremic toxins that are able to elicit protein damage, guanidine produced a dose-dependent increase in protein damage. No difference was found after a hemodialysis session. Deamidated albumin shows normal binding capacity to warfarin, salicylic acid, or diazepam but reduced binding to homocysteine. In conclusion, uremic toxins, especially guanidine, display an ability to induce significant protein damage, which can in turn have functional consequences. Copyright & 2004 by the American Society of Nephrology.

Purpose of reviewThis review focuses on recent findings (June 2002-July 2003) on the topic of homocysteine, a sulfur amino acid associated with cardiovascular disease, and its metabolism in renal failure, a condition with a high prevalence of both hyperhomocysteinemia and cardiovascular disease.Recent findingsA large meta-analysis of prospective studies in the general population established that hyperhomocysteinemia is a risk factor for cardiovascular disease. The results of intervention trials, once available, will also have to be tested in a meta-analysis, because of predicted problems with their statistical power. In kidney patients, intervention trials, still in the recruiting stage, target transplant patients, because of their unique characteristics related to folate responsiveness. As for the cause of hyperhomocysteinemia, new findings show that in humans, renal metabolic extraction depends on renal plasma flow in the post-absorptive state. Folate absorption or interconversion seems not to be affected. Riboflavin is a determinant of plasma homocysteine levels in uraemia. The consequences of hyperhomocysteinemia in uraemia are DNA hypomethylation and altered gene expression.SummaryThe causes of hyperhomocysteinemia in renal failure are still not clear. However, the possibilities include defective renal or extrarenal metabolism as a result of uraemic toxicity. Renal plasma flow is important in homocysteine renal metabolism. Among the consequences of hyperhomocysteinemia in renal failure are impaired protein and DNA methylation, with an alteration in the allelic expression of genes regulated through methylation. Intervention trials are under way to test whether hyperhomocysteinemia is causally related to cardiovascular disease in this patient population.

BackgroundBed-rest experiments are designed for investigation on catabolic effects of hypokinetic conditions and/or for microgravity simulation in on-ground aerospace research. Bed-rest effects include a reduction in fat-free mass and muscle mass. Urea and creatinine are catabolites of endogenous protein and of muscular energetic metabolism which are excreted mainly by the kidney. The study investigated on urea, creatinine, and kidney function during bed-rest.MethodsTwenty healthy young men underwent a 7-day adaptation period (day-6 to day-0) and a 35-day bed-rest experiment (day1 to day35) during normocaloric diet. Urine were collected from day-3 to day0 (baseline) and from day1 to day35. Blood samples and anthropometrical data were collected at day0 (baseline) and bed-rest days 7, 14, 21, 28, and 35.ResultsBed-rest reduced plasma volume, weight, fat-free mass, and muscle mass (PConclusionsStudy results shows that urea is a marker of catabolic conditions secondary to hypokinetic conditions.

The study analyzes the prevalence of cardiovascular risk factors in 1343 patients with severe headache (399 men and 944 women), aged 15 to 64 years; analyses were controlled for sex, age, and type and frequency of headache. Prevalence of various forms of headache was different between men and women. Age and days per year with headache were significantly different among various forms of headache. For men and women with headache, age directly related to prevalence of hypertension, hypercholesterolemia, and obesity. Due to low prevalence, analyses by age were not done for diabetes mellitus. For cigarette smoking, prevalence was not related to age in men, but was inversely related to age in women. With control for age, prevalence of cardiovascular risk factors was not significantly different among patients with different forms of headache, except for cluster headache. Among men with cluster headache, prevalence was high for cigarette smoking, but low for hypercholesterolemia. With control for age, days per year with headache did not relate to prevalence of cardiovascular risk factors except for cigarette smoking in men. Compared to data for a population sample used as control, patients with headache had higher prevalence of hypertension in both sexes, independent of age (odds ratio 1.51, 95% confidence interval 1.28 to 1.80); the difference between patients with headache and the control population was lower with increasing age. The high prevalence of hypertension among patients with headache was not due to overweight. The data indicate that headache is significantly associated with hypertension, but not with other cardiovascular risk factors.

The cardiovascular risk factor homocysteine is mainly bound to proteins in human plasma, and it has been hypothesized that homocysteinylated proteins are important mediators of the toxic effects of hyperhomocysteinemia. It has been recently demonstrated that homocysteinylated proteins are elevated in hemodialysis patients, a high cardiovascular risk population, and that homocysteinylated albumin shows altered properties. OBJECTIVE: Aim of this work was to investigate the effects of homocysteinylated albumin - the circulating form of this amino acid, utilized at the concentration present in uremia - on monocyte adhesion to a human endothelial cell culture monolayer and the relevant molecular changes induced at both cell levels. METHODS AND RESULTS: Treated endothelial cells showed a significant increase in monocyte adhesion. Endothelial cells showed after treatment a significant, specific and time-dependent increase in ICAM1 and VCAM1. Expression profiling and real time PCR, as well as protein analysis, showed an increase in the expression of genes encoding for chemokines/cytokines regulating the adhesion process and mediators of vascular remodeling (ADAM17, MCP1, and Hsp60). The mature form of ADAM17 was also increased as well as Tnf-α released in the cell medium. At monocyte level, treatment induced up-regulation of ICAM1, MCP1 and its receptor CCR2. CONCLUSIONS: Treatment with homocysteinylated albumin specifically increases monocyte adhesion to endothelial cells through up-regulation of effectors involved in vascular remodeling. Rationale: The cardiovascular risk factor homocysteine is mainly bound to proteins in human plasma, and it has been hypothesized that homocysteinylated proteins are important mediators of the toxic effects of hyperhomocysteinemia. It has been recently demonstrated that homocysteinylated proteins are elevated in hemodialysis patients, a high cardiovascular risk population, and that homocysteinylated albumin shows altered properties. Objective: Aim of this work was to investigate the effects of homocysteinylated albumin - the circulating form of this amino acid, utilized at the concentration present in uremia - on monocyte adhesion to a human endothelial cell culture monolayer and the relevant molecular changes induced at both cell levels. Methods and Results: Treated endothelial cells showed a significant increase in monocyte adhesion. Endothelial cells showed after treatment a significant, specific and time-dependent increase in ICAM1 and VCAM1. Expression profiling and real time PCR, as well as protein analysis, showed an increase in the expression of genes encoding for chemokines/cytokines regulating the adhesion process and mediators of vascular remodeling (ADAM17, MCP1, and Hsp60). The mature form of ADAM17 was also increased as well as Tnf-α released in the cell medium. At monocyte level, treatment induced up-regulation of ICAM1, MCP1 and its receptor CCR2. Conclusions: Treatment with homocysteinylated albumin specifically increases monocyte adhesion to endothelial cells through up-regulation of effectors involved in vascular remodeling. © 2012 Capasso et al.

Introduction: The Kidney Disease: Improving Global Outcomes (KDIGO) foundation promoted the establishment of the Chronic Kidney Disease (CKD) Prognosis Consortium to meta-analyze the association of estimated glomerular filtration rate (eGFR) and albuminuria with incidence of various outcomes in samples of general populations from all over the world. Methods: Variables in meta-analysis included eGFR by the Modification of Diet in Renal Disease (MDRD) Study equation, the urinary albumin to creatinine ratio (uACR) as index of albuminuria, together with proteinuria at dipstick urinalysis and classical markers of cardiovascular risk. Overall, 105,872 participants had uACR measurements, and 1,128,310 participants had dipstick measurements. Results: The association with mortality was continuous over the whole range of uACR/proteinuria and Jshaped for eGFR which was associated with an excess risk for values

Background. Hydrogen sulphide, H2S, is the third endogenous gas with putative cardiovascular properties, after nitric oxide and carbon monoxide. H2S is a vasorelaxant, while H2S deficiency is implicated in the pathogenesis of hypertension and atherosclerosis. Cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE) and 3-mercaptopyruvate sulphurtransferase (MPS) catalyze H2S formation, with different relative efficiencies. Chronic kidney disease (CKD) is characterized by elevation of both plasma homocysteine and cysteine, which are substrates of these enzymes, and by a high prevalence of hypertension and cardiovascular mortality, particularly in the haemodialysis stage. It is possible that the H2S-generating pathways are altered as well in this patient population. Methods. Plasma H2S levels were measured with a common spectrophotometric method. This method detects various forms of H2S, protein-bound and non-protein-bound. Blood sulphaemoglobin, a marker of chronic exposure to H2S, was also measured, as well as related sulphur amino acids, vitamins and transcriptional levels of relevant genes, in haemodialysis patients and compared to healthy controls. Results. Applying the above-mentioned methodology, H2S levels were found to be decreased in patients. Sulphaemoglobin levels were significantly lower as well. Plasma homocysteine and cysteine were significantly higher; vitamin B6, a cofactor in H2S biosynthesis, was not different. H2S correlated negatively with cysteine levels. CSE expression was significantly downregulated in haemodialysis patients. Conclusions. Transcriptional deregulation of genes encoding for H2S-producing enzymes is present in uraemia. Although the specificity of the method employed for H2S detection is low, the finding that H2S is decreased is complemented by the lower sulphhaemoglobin levels. Potential implications of this study relate to the pathogenesis of the uraemic syndromemanifestations, such as hypertension and atherosclerosis. Background. Hydrogen sulphide, H2S, is the third endogenous gas with putative cardiovascular properties, after nitric oxide and carbon monoxide. H2S is a vasorelaxant, while H2S deficiency is implicated in the pathogenesis of hypertension and atherosclerosis. Cystathionine beta-synthase (CBS), cystathionine gamma-lyase (CSE) and 3-mercaptopyruvate sulphurtransferase (MPS) catalyze H2S formation, with different relative efficiencies. Chronic kidney disease (CKD) is characterized by elevation of both plasma homocysteine and cysteine, which are substrates of these enzymes, and by a high prevalence of hypertension and cardiovascular mortality, particularly in the haemodialysis stage. It is possible that the H2S-generating pathways are altered as well in this patient population.Methods. Plasma H2S levels were measured with a common spectrophotometric method. This method detects various forms of H2S, protein-bound and non-protein-bound. Blood sulphaemoglobin, a marker of chronic exposure to H2S, was also measured, as well as related sulphur amino acids, vitamins and transcriptional levels of relevant genes, in haemodialysis patients and compared to healthy controls.Results. Applying the above-mentioned methodology, H2S levels were found to be decreased in patients. Sulphaemoglobin levels were significantly lower as well. Plasma homocysteine and cysteine were significantly higher; vitamin B-6, a cofactor in H2S biosynthesis, was not different. H2S correlated negatively with cysteine levels. CSE expression was significantly downregulated in haemodialysis patients.Conclusions. Transcriptional deregulation of genes encoding for H2S-producing enzymes is present in uraemia. Although the specificity of the method employed for H2S detection is low, the finding that H2S is decreased is complemented by the lower sulphhaemoglobin levels. Potential implications of this study relate to the pathogenesis of the uraemic syndrome manifestations, such as hypertension and atherosclerosis.

High levels of homocysteine have been implicated as a cardiovascular risk factor in the general population and in patients with chronic renal failure, and particularly patients on hemodialysis. To classify a risk factor as causally related to a certain disease, both strong epidemiologic data and sound basic-science studies establishing a mechanism are needed. Among the latter, the hypomethylation of proteins and DNA, and protein homocysteinylation, have been investigated in uremia, providing for an array of toxic effects in this disease. © 2008 National Kidney Foundation, Inc.

Hyperhomocysteinemia, highly prevalent in well-nourished patients with chronic renal failure and in uremia, causes toxic effects that can be envisioned in terms of cardiovascular risk increase. However, its effects on cellular metabolism and on gene expression, not to mention receptor regulation, only recently are being evaluated. For example, it has been shown that hypomethylation induced by hyperhomocysteinemia can alter erythrocyte membrane protein repair and gene expression. In addition, increased plasma protein L-isoaspartyl content, related to hyperhomocysteinemia and uremic toxicity, determines specific effects on protein function, with a reduced binding of homocysteine to albumin. We propose that uremia is a state in which proteins present a widespread derangement of structure-function relationships. Hyperhomocysteinemia, highly prevalent in well-nourished patients with chronic renal failure and in uremia, causes toxic effects that can be envisioned in terms of cardiovascular risk increase. However, its effects on cellular metabolism and on gene expression, not to mention receptor regulation, only recently are being evaluated. For example, it has been shown that hypomethylation induced by hyperhomocysteinemia can alter erythrocyte membrane protein repair and gene expression. In addition, increased plasma protein L-isoaspartyl content, related to hyperhomocysteinemia and uremic toxicity, determines specific effects on protein function, with a reduced binding of homocysteine to albumin. We propose that uremia is a state in which proteins present a widespread derangement of structure-function relationships. © 2006 Elsevier Inc. All rights reserved.

Estimated glomerular filtration rate (eGFR) was used for analysis of kidney disease prevalence in the United States. The study investigated on prevalence, associated disorders, and kidney disease awareness in an Italian population sample. Data were collected on serum creatinine, other laboratory indices, blood pressure, and medical history in the Gubbio Population Study (n=4574, both sexes, ages 18-95 years). Analyses were carried out on eGFR (equation of Modification Diet in Renal Disease study), disorders potentially secondary to kidney dysfunction (hypertension, high serum uric acid, high serum phosphorus/low serum calcium, high serum potassium, cardiovascular disease, anemia), and kidney disease awareness. The prevalence of eGFR60 ml/min x 1.73 m(2) increased with age in both sexes (from1% for ages 18-24 years to30% for agesor =75 years, P0.001). In the group with eGFR60 ml/min x 1.73 m(2), number of disorders secondary to kidney dysfunction wasor =2 in the majority of persons, was higher than in persons with eGFRor =60 ml/min x 1.73 m(2) (P0.001), and was inversely related to eGFR (P0.001). The prevalence of reported kidney disease was 3.3% in the group with eGFR60 ml/min x 1.73 m(2) and directly related to serum creatinine and number of disorders secondary to kidney dysfunction (P0.001). Low kidney function is frequent in the older population and is associated with disorders typical of kidney disease. Persons with low kidney function are rarely aware of kidney disease unless of very high serum creatinine or presence of many disorders typical of kidney disease.

Hyperhomocysteinemia, an independent cardiovascular risk factor, is present in the majority of hemodialysis patients. Among the postulated mechanisms of toxicity, protein homocysteinylation is potentially able to cause significant alterations in protein function. Protein homocysteinylation occurs through various mechanisms, among which is the post-translational acylation of free amino groups (protein-Nhomocysteinylation, mediated by homocysteine (Hcy) thiolactone). Another type of protein homocysteinylation occurs through the formation of a covalent *S*S* bond, found primarily with cysteine residues (protein-S-homocysteinylation). Scant data are available in the literature regarding the extent to which alterations in protein homocysteinylation are present in uremic patients on hemodialysis, and the effects of folate treatment are not known. Protein homocysteinylation was measured in a group of hemodialysis patients (n¼28) compared to controls (n¼14), with a new method combining protein reduction, gel filtration and Hcy derivatization. Chemical hydrolysis was performed, followed by highpressure liquid chromatography separation. The effects of folate treatment on protein homocysteinylation, as well as in vitro binding characteristics were evaluated. Plasma Hcy, protein-N-homocysteinylation and protein-S-homocysteinylation were significantly higher in patients vs controls. Plasma Hcy and protein-S-homocysteinylation were significantly correlated. After 2 months of oral folate treatment, protein-Nhomocysteinylation was normalized, and protein-S-homocysteinylation was significantly reduced. Studies on albuminbinding capacity after in vitro homocysteinylation show that homocysteinylated albumin is significantly altered at the diazepam-binding site. In conclusion, increased protein homocysteinylation is present in hemodialysis patients, with possible consequences in terms of protein function. This alteration can be partially reversed after folate treatment. Hyperhomocysteinemia, an independent cardiovascular risk factor, is present in the majority of hemodialysis patients. Among the postulated mechanisms of toxicity, protein homocysteinylation is potentially able to cause significant alterations in protein function. Protein homocysteinylation occurs through various mechanisms, among which is the post-translational acylation of free amino groups (protein-N-homocysteinylation, mediated by homocysteine (Hcy) thiolactone). Another type of protein homocysteinylation occurs through the formation of a covalent -S-S- bond, found primarily with cysteine residues (protein-S-homocysteinylation). Scant data are available in the literature regarding the extent to which alterations in protein homocysteinylation are present in uremic patients on hemodialysis, and the effects of folate treatment are not known. Protein homocysteinylation was measured in a group of hemodialysis patients (n = 28) compared to controls (n = 14), with a new method combining protein reduction, gel filtration and Hcy derivatization. Chemical hydrolysis was performed, followed by high-pressure liquid chromatography separation. The effects of folate treatment on protein homocysteinylation, as well as in vitro binding characteristics were evaluated. Plasma Hcy, protein-N-homocysteinylation and protein-S-homocysteinylation were significantly higher in patients vs controls. Plasma Hcy and protein-S-homocysteinylation were significantly correlated. After 2 months of oral folate treatment, protein-N-homocysteinylation was normalized, and protein-S-homocysteinylation was significantly reduced. Studies on albumin-binding capacity after in vitro homocysteinylation show that homocysteinylated albumin is significantly altered at the diazepam-binding site. In conclusion, increased protein homocysteinylation is present in hemodialysis patients, with possible consequences in terms of protein function. This alteration can be partially reversed after folate treatment. © 2006 International Society of Nephrology.

Fish-odor syndrome or trimethylaminuria (OMIM #602079), is a rare inborn error of metabolism inherited in an autosomal recessive fashion, involving the dysfunction of hepatic enzyme flavin-containing monooxygenase 3 (FMO3) that converts fishy-smelling trimethylamine (TMA) into odorless trimethylamine-N-oxide (TMAO). This confers, to the affected individual a very unpleasant body odor resembling that of rotting fish. This disorder has been relatively well-documented in British, Australian, and American populations and reports have appeared regarding patients in Thailand and Hong Kong, but no Italian families affected by trimethylaminuria have been reported in the literature. We have collected a cohort of Italian families and investigated the genetic basis of the disorder in these Italian pedigrees disclosing a spectrum of molecular variation in the FM03 gene comprising three novel deleterious mutations: the first documented de novo missense mutation causative of trimethylaminuria; a guanidine nucleotide deletion (G1182del) at codon 394 and a novel missense mutation (R238P) that altered highly conserved amino acid in the exon 6. Moreover, we investigated by aplotype analysis a family with mild TMAuria identifying a putative causative aplotype. Finally, we failed to detect any variation in other Italian families suggesting that this gene is not associated with all clinical form of trimethylaminuria or that polymorphisms in this gene could be susceptibility factors for developing the disease. Our findings support the hypothesis that TMAuria is not a rare recessive disorder but rather a spectrum of malodour phenotypes in which diet and environmental exposures can play a role in triggering symptoms.

In uremia, plasma levels of anti-protein C and anti-protein S antibodies are associated with thrombosis. Background Vascular access thrombosis is an important cause of morbidity in patients with end-stage renal failure on maintenance hemodialysis (MHD). However, little is known about its risk factors. The present study was undertaken to evaluate the role of coagulation factors, fibrinolytic factors, and anti-phospholipid antibodies (aPL). In particular, we have evaluated the role of anti-protein C and anti-protein S antibodies in patients on MHD with and without thrombosis because no data are available in the literature. Methods The study group comprised 30 patients with thrombotic complications (TC), 40 patients matched for age, sex, and dialytic age with no thrombotic complications (NTC) and 400 controls. We have measured: anti-protein C antibodies, anti-protein S antibodies, anticardiolipin antibodies (ACA), anti-β2-glycoprotein antibodies (β2-GPI), and anti-prothrombin antibodies (aPT), along with prothrombin time, fibrinogen, plasminogen, protein C, protein S, anti-thrombin III, APC-resistance test, D-dimer, tissue-type plasminogen's activator, plasminogen activator inhibitor-1 (PAI-1), prothrombin fragment 1+2, factors of the intrinsic and extrinsic pathway, C-reactive protein, and homocysteine. Results There were no significant differences between groups for prothrombin time, fibrinogen, plasminogen, protein C, protein S, anti-thrombin III, activated protein C (APC) resistance, D-dimer, tPA, C-reactive protein, Factors II, X, and VII. The anti-β2-GP1 and aPT were elevated in both TC and NTC patients, compared to the control group. Significant differences between TC and NTC groups were found for anti-protein C and anti-protein S antibodies, ACA-IgM, PAI-1, Factor VIII, prothrombin fragments 1+2, and homocysteine. Conclusion The most novel finding was a significant elevation of anti-protein C antibodies and anti-protein S antibodies in the TC group (i.e., in patients on MHD with thrombosis of vascular access). It indicates that other pathogenetic mechanisms in addition to endothelial damage may cause hypercoagulability in uremia.

The moderate increase in urinary albumin excretion defined as microalbuminuria is not rare and is associated with cardiovascular risk factors. Microalbuminuria prevalence is low in the absence of cardiovascular risk factors and progressively increases with the number cardiovascular risk factors. The main correlate of microalbuminuria is blood pressure, either systolic or diastolic pressure. The relation between blood pressure and microalbuminuria is continuous and graded because the microalbuminuria prevalence increases with the severity of hypertension. Among hypertensive patients on drug treatment, blood pressure control is associated with a low prevalence of microalbuminuria. Thus, blood pressure appears as a determinant of microalbuminuria rather than a mere correlate. For hypercholesterolemia, smoking, and diabetes, data are less strong but point to an independent positive association with microalbuminuria. Altogether, data indicate that microalbuminuria in the population reflects the presence of cardiovascular risk factors. Data on microalbuminuria and coronary heart disease support this idea. There is a continuous and graded relation between urinary albumin excretion and coronary heart disease prevalence. High urinary albumin excretion is likely a sign of vascular damage existing both at the renal and cardiac levels and induced by 1 or more uncontrolled cardiovascular risk factors.

Hyperhomocysteinemia is present in the majority of well-nourished chronic renal failure and uremic patients. Most observations reported in the literature come from studies carried out in end-stage renal disease patients treated with hemodialysis. The underlying mechanisms of the toxic effects of homocysteine in uremia related to cardiovascular disease and other disturbances are still under scrutiny. As a consequence, macromolecules (i.e., proteins and DNA) have been found to be altered to various extents. One of the mechanisms of homocysteine toxicity is related to the action of its metabolic precursor, S-adenosylhomocysteine, a powerful methyltransferase competitive inhibitor. Disruption of DNA methylation has been demonstrated to occur as a result of hyperhomocysteinemia, and/or is associated with vascular damage. DNA hypomethylation has been found in the mononuclear cell fraction of uremic patients with hyperhomocysteinemia. Proteins are also targets of homocysteine-dependent molecular damage. The formation of oxidative products with free cysteinyl residue thiol groups has been demonstrated to occur in blood. The latter also represents a mechanism for the transport of homocysteine in plasma. In addition, homocysteine thiolactone has been shown to react with free amino groups in proteins to form isopeptide bonds, in particular at the lysine residue level. Another type of isopeptide bond in proteins may result from the deamidation and isomerization of asparaginyl residues, yielding abnormal isoaspartyl residues, which have been demonstrated to be increased in uremic patients. Folate treatment exerts a partial, but significant, homocysteine-lowering effect in uremic patients and has been shown to improve the changes in macromolecules induced by high homocysteine levels. In conclusion, both DNA and proteins are structurally modified in uremia as a consequence of high homocysteine levels. The role of these macromolecule changes in inducing the clinical complications of hyperhomocysteinemia in these patients, although still conjectural in some respects, is at present sustained by several pieces of evidence.

In the general population, hyperhomocysteinemia is an independent risk factor for cardiovascular disease (ischemic disease, such as stroke and myocardial infarction, and arterial and venous thrombosis). We can presume that the association is causal, based on the example of homocystinuria, and on the evidence put forward by several basic science and epidemiologic studies. However, the results of large intervention trials, which may grant further support to this hypothesis, are not yet available. In chronic renal failure and in uremia, the evidence that is offered by carefully performed prospective studies also indicate the presence of an association, although some studies suggest reverse epidemiology. The mechanisms underlying the association, and able to explain the several toxic effects of homocysteine, related or not to cardiovascular disease, are unclear. Oxidation, nitrosylation, and hypomethylation are among the postulated mechanisms. In uremia, protein hypomethylation interferes with protein repair; DNA hypomethylation impairs regulation of gene expression, whereas folate treatment reverts such alterations. Acylation, another structural modification able to impair protein function, is a possible mediator of homocysteine toxicity. © 2004 Elsevier Inc. All rights reserved.

Possible mechanisms of homocysteine toxicity. Hyperhomocysteinemia is a risk factor for cardiovascular disease in the general population. In chronic renal failure (CRF), plasma homocysteine levels rise when the glomerular filtration rate (GFR) is reduced 50%, and in uremia the majority of patients are hyperhomocysteinemic. The purpose of this study was to review possible mechanisms of homocysteine toxicity. Homocysteine, a sulfur amino acid found in blood in micromolar concentrations, can have toxic effects through a handful of general possible mechanisms. These mechanisms include oxidative stress (through the production of reactive oxygen species), binding to nitric oxide, production of homocysteinylated/acylated proteins, and accumulation of its precursor, S-adenosyl-homocysteine, a potent inhibitor of transmethylation reactions. Methyltransferase inhibition actually occurs in CRF and in uremia, and can have several functional consequences.

Recently, we have measured in erythrocytes a voltage-modulated and dihydropyridine-inhibited calcium influx. Since arachidonic acid and other polyunsaturated fatty acids influence the activities of most ion channels, we studied their effects on the erythrocyte Ca(2+) influx. It was measured on fresh erythrocytes, isolated from healthy donors, using the fluorescent dye Fura 2 as indicator of [Ca(2+)](i). AA (5-50 microM) and EPA (20-30 microM) stimulated a concentration-dependent increase in [Ca(2+)](i), deriving from extracellular calcium (1 mM), without affecting the intra- and extracellular pH and membrane voltage. The Ca(2+) influx rate varied from 0.5 to 3 nM Ca(2+)/s in the presence of AA and from 0.9 to 1.7 nM Ca(2+)/s with EPA. The Ca(2+) influx elicited by AA and EPA was not inhibited by dihydropyridines, while cyclooxygenase inhibitors were effective and PGE1 or PGE2 did not produce any effect. We conclude that AA could activate an erythrocyte voltage-independent Ca(2+) transport via an intermediate product of cyclooxygenase pathway; however, a direct interaction with the membrane lipid-protein cannot be excluded.

A previous study reported that urinary markers of protein intake are inversely related to blood pressure via unknown mechanisms. In man and rats, protein intake affects renal function and increases renal sodium excretion. The present study investigates the relation between markers of protein intake and blood pressure and the possible role of sodium in this relation. Blood pressure status, overnight urinary urea as index of protein intake, urinary and plasma sodium, and other variables were measured in a population sample of 3705 men and women, aged 25–74 years, without high plasma creatinine. Urinary urea was inversely related to blood pressure and hypertension: in multivariate analyses, 6.5 mmol/h higher urinary urea (about one s.d. in men and women) was related to 4.25 mm Hg lower systolic blood pressure (95% confidence interval = 1.34–8.49), and to 0.65 lower risk of hypertension (95% CI 0.34–0.87). An interaction was found between overnight urinary sodium and the relation of urinary urea to blood pressure: the relation was significant only in persons with overnight urinary sodium above the median. Urinary urea was significantly and inversely also related to plasma sodium. Data confirm an inverse relation to blood pressure of protein intake as measured by urinary urea. The possibility of sodium-related mechanisms is supported by the interaction of urinary sodium with the relation and by the inverse association of urinary urea with plasma sodium. The hypothesis is made that high protein intake could counteract sodium-dependent blood pressure rise via stimulation of renal sodium excretion.

Hyperhomocysteinemia is an independent cardiovascular risk factor that possibly accounts for about one of 5 cardiovascular deaths. It is conceivable that the importance of hyperhomocysteinemia will increase when other risk factors, such as hypertension or hypercholesterolemia, will become less prevalent in the general population. In chronic renal failure (CRF), high plasma homocysteine levels are a common finding and in uremia almost the rule. However, a small subset of patients remains normohomocysteinemic. The cause of hyperhomocysteinemia in CRF, whether it lies in an impaired renal or extrarenal metabolism or through uremic retention toxins, is still under intensive scrutiny. As for the consequences of high homocysteine levels in the general population and in patients with CRF, these are many-fold and linked to the mechanism of homocysteine toxic action. In fact, homocysteine can be harmful to cells because (1) it evokes oxidative stress (through the production of reactive oxygen species), (2) binds to nitric oxide, (3) produces homocysteinylated proteins, or (4) leads to the accumulation of its precursor, S-adenosylhomocysteine, a potent inhibitor of biological transmethylations. Macromolecule hypomethylation is a common feature in CRF and uremia with possible functional consequences. Nutritional or pharmacologic interventions have been proposed in the treatment of hyperhomocysteinemia, while the results of large clinical trials designed to assess if lowering homocysteine levels is effective in reducing cardiovascular risk, are pending. © 2003 by the National Kidney Foundation, Inc.

Abstract —The basic requirement for declaring an association study positive is that the “hypertension-favoring” allele is more frequent in hypertensive cases than in normotensive controls. However, both positive and negative associations with hypertension have been found for the same polymorphism when studied in different populations. In the present study, we addressed the question of the possible cause(s) of this discrepancy among populations by using the α-adducin polymorphism as a paradigm. Four hundred ninety hypertensives and 176 normotensives enrolled in Sassari, Italy, and 468 hypertensives and 181 normotensives enrolled in Milano, Italy, were genotyped for the α-adducin Gly460Trp polymorphism. The blood pressure response to 2 months of hydrochlorothiazide therapy could be evaluated in 143 (85 in Sassari and 58 in Milano) hypertensives with and without the 460Trp α-adducin allele. The α-adducin 460Trp allele was not significantly more frequent in hypertensives in the Sassari population but was more frequent in hypertensives than in normotensives in Milano ( P =0.019). Basal plasma renin activity was lower and blood pressure fall after diuretic therapy more pronounced ( P