Conceptual basis and methodology of the SOPHIA study.

To clarify the role of gene polymorphisms on the effect of losartan and losartan plus hydrochlorothiazide on blood pressure (primary end point) and on cardiac, vascular and metabolic phenotypes (secondary end point) after 4, 8, 12, 16 and 48 weeks treatment, an Italian collaborative study - The Study of the Pharmacogenomics in Italian hypertensive patients treated with the Angiotensin receptor blocker losartan (SOPHIA) - on never-treated essential hypertensives (n = 800) was planned. After an 8 week run-in, losartan 50 mg once daily will be given and doubled to 100 mg at week +4 if blood pressure is more than 140/90 mmHg. Hydroclorothiazide 25 mg once daily at week +8 and amlodipine 5 mg at week +16 will be added if blood pressure is more than 140/90 mmHg. Cardiac mass (echocardiography), carotid intima-media thickness, 24 h ambulatory blood pressure, homeostatic model assessment (HOMA) index, microalbuminuria, plasma renin activity and aldosterone, endogenous lithium clearance, brain natriuretic peptide and losartan metabolites will be evaluated. Genes of the renin-angiotensin-aldosterone system, salt sensitivity, the beta-adrenergic system and losartan metabolism will be studied (Illumina custom arrays). A whole-genome scan will also be performed in half of the study cohort (1M array, Illumina 500 GX beadstation).

Fetal ontogeny and role of metanephric bradykinin B2 receptors.

Previous studies in rats have shown that blockade of bradykinin B2 receptors (B2R) in combination with a high-salt intake during gestation result in poor postnatal survival and long-term hypertension in the offspring. In this study, we examined the fetal ontogeny of B2R and determined the consequences of gestational B2R blockade and high salt on kidney development. B2R gene expression is induced on embryonic day (E16) of fetal metanephrogenesis and remains sustained until term. The earliest expression of the B2R protein is observed on apical membranes of ureteric bud branches and in capillary loop stage glomeruli. By the end of gestation, B2R becomes restricted to more-differentiated tubules in the deep cortex and medulla. Pairs of rats on normal (0.12 mmol/g) or high (0.84 mmol/g) salt diets were mated at 14 weeks of age. The B2R antagonist, Icatibant (previously known as Hoe-140) (300 nmol/kg per day) or saline (vehicle) was infused intraperitoneally during gestation via osmotic minipumps. Fetuses were examined on E20 (n=27-36 per group). No significant differences in litter size or body weight were observed among the groups. Combined high-salt and Icatibant treatment caused aberrant fetal renal development characterized by tubular dysgenesis, widened stromal mesenchyme, and glomerular cysts. The dysgenetic tubules stained positively for the distal nephron lectin, Dolichos biflorus, and exhibited enhanced Bax expression and apoptosis. Renal microvascular development, the number of mature glomeruli, and percentage of proliferating glomerular cells were not affected. Gestational Icatibant or high salt alone had no deleterious effects on fetal nephrogenesis. We conclude that gestational blockade of the kallikrein-kinin system impairs fetal nephrogenesis if combined with an intrauterine stressor such as high-salt intake. B2R may play a protective role during segmental nephron differentiation.

High plasma prorenin in non diabetic siblings of non insulin-dependent diabetes mellitus patients.

In a large cohort (no. = 361) of NIDDM probands and their concordant/discordant siblings from no. = 132 families we studied: 1. the levels of plasma prorenin in non affected siblings of NIDDM probands as opposed to normal subjects without family history of diabetes, and 2. whether plasma prorenin raises in parallel to urinary protein loss in NIDDM patients. Prorenin (solid-phase trypsin) and micro-macroalbuminuria (radioimmunoassay) were evaluated. Plasma prorenin was higher in NIDDM probands and siblings than in non NIDDM siblings (37+/-31 vs. 25+/-15 ng/ml/h, p<0.0005) who, in turn, showed higher plasma prorenin than non diabetic controls without family history of diabetes (25+/-15 vs. 17+/-8 ng/ml/h, p<0.005). Plasma prorenin was higher in NIDDM siblings of micro-macroalbuminuric probands than in NIDDM siblings of non micro-macroalbuminuric probands (40+/-26 vs. 29+/-20 ng/ml/h, mean +/- SD, p = 0.0058) whereas no difference was found among non diabetic siblings (24+/-14 vs. 22+/-11 ng/ml/h, NS). Our data confirm that plasma prorenin is elevated in NIDDM patients, and show: 1. that the raise of plasma prorenin in non-NIDDM siblings of a diabetic patient does not depend entirely from the presence of diabetes, and 2. that plasma prorenin in NIDDM probands and their concordant siblings goes along with micro-macroalbuminuria.

Blood pressure sensitivity to salt in rats with low urinary kallikrein excretion.

We evaluated if a rat strain inbred for reduced urinary kallikrein excretion differs from normal-kallikrein Wistar rats regarding blood pressure in basal conditions and during alterations in sodium balance. Low-kallikrein rats showed greater systolic blood pressure values (125 +/- 3 vs. 114 +/- 2 mmHg in controls, P < 0.01) at 9 weeks of age. Systolic blood pressure was increased after 20 days of dietary sodium loading in the low-kallikrein group and remained unchanged in controls (150 +/- 6 vs. 112 +/- 2 mmHg, P < 0.01) and this effect was associated with a reduced cumulative excretion of sodium (23% less in the low-kallikrein group compared with controls, P < 0.01). Urinary creatinine excretion was decreased by sodium loading in both groups, and this effect was more pronounced in the low-kallikrein group. The group-difference in urinary kallikrein excretion found in basal conditions (2.49 +/- 0.10 vs. 7.78 +/- 0.53 Pkat/100 g body weight, P < 0.01) was enhanced by high salt diet (1.05 +/- 0.21 vs. 8.31 +/- 0.70 Pkat/100 g body weight, P < 0.01). The ratio of heart weight to body weight was significantly greater in low-kallikrein rats (331 +/- 7 vs. 275 +/- 4 mg/100 g body weight, P < 0.01), whereas the ratio of kidney weight to body weight was lower (329 +/- 5 vs. 370 +/- 8 mg/100 body weight, P < 0.01). Our results indicate that a genetically-determined defect in urinary kallikrein excretion is associated with a greater blood pressure sensitivity to salt, possibly due to altered renal sodium handling.

Urinary kallikrein: a marker of blood pressure sensitivity to salt.

We evaluated if a rat strain inbred for low urinary kallikrein excretion differs from normal-kallikrein Wistar rats regarding blood pressure levels in basal conditions and during alterations in sodium balance. Blood pressure was measured in unanesthetized rats on normal sodium intake. Then, blood pressure sensitivity to salt was evaluated over a period of 20 days of high sodium diet (0.84 mmol per g chow). Low-kallikrein rats showed greater systolic blood pressure levels (125 +/- 3 vs. 114 +/- 2 mm Hg in controls, P < 0.01) at nine weeks of age. Systolic blood pressure was increased after sodium loading in the low-kallikrein group and remained unchanged in controls (150 +/- 6 vs. 112 +/- 2 mm Hg, P < 0.01). This effect was associated with a reduced cumulative urinary excretion of sodium in the low-kallikrein rats. No group difference was found in the clearance of endogenous creatinine in basal conditions. Urinary creatinine excretion decreased during sodium loading, particularly in the low-kallikrein group. The group-difference in urinary kallikrein excretion found in basal conditions (6.85 +/- 0.31 vs. 20.74 +/- 1.71 nkat/24 hr in controls, P < 0.01) was enhanced by high salt diet (2.96 +/- 0.67 vs. 22.07 +/- 2.47 nkat/24 hr in controls, P < 0.01). In addition, renal kallikrein activity and content were reduced in low-kallikrein rats. The latter group showed a greater ratio of heart weight to body wt both in basal conditions and after sodium loading. The ratio of kidney weight to body wt was reduced after sodium loading. These results indicate that a genetically-determined defect in urinary kallikrein excretion is associated with a greater blood pressure sensitivity to salt, possibly due to altered renal sodium handling.

Sexual dimorphism of cardiovascular responses to early blockade of bradykinin receptors.

To assess whether the cardiovascular effects induced by early blockade of bradykinin B2-receptors with Hoe 140 (D-Arg[Hyp3,Thi5,D-Tic7,Oic8]-bradykinin) are influenced by sex, Wistar rats of both sexes received the antagonist (300 nmol/d per kilogram body wt) or vehicle from 2 days to 7 weeks of age by subcutaneous injection and then by intraperitoneal infusion. Compared with control rats, Hoe 140-treated female rats showed higher systolic blood pressure levels at 7 and 9 weeks of age (125 +/- 2 versus 111 +/- 2 mm Hg and 132 +/- 3 versus 116 +/- 2 mm Hg, respectively, P < .05), whereas in male rats a difference was found at 7 weeks (122 +/- 4 versus 108 +/- 4 mm Hg, P < .05) but not at 9 weeks. At this stage, the mean blood pressure of Hoe 140-treated rats was higher than that of control animals, and this difference was more pronounced at 12 weeks in female rats (121 +/- 2 versus 100 +/- 3 mm Hg in control animals, P < .01) compared with males (116 +/- 3 versus 104 +/- 2 mm Hg in control animals, P < .05). After the first week of life, body weight gain was greater in Hoe 140-treated female rats than in control rats, whereas a group-difference was detected in male rats only after weaning. In Hoe 140-treated female rats, heart weight was already increased at 9 weeks (330 +/- 6 versus 305 +/- 5 mg/100 g body wt in control rats, P < .05), whereas it was necessary to prolong Hoe 140 administration in male rats to develop heart hypertrophy (300 +/- 4 versus 275 +/- 4 mg/100 g body wt in control rats at 12 weeks, P < .05). Tissue kallikrein mRNA levels were higher in the kidney of adult female rats, whereas no sex difference was detected in the heart. The finding of a sexual dimorphism in the cardiovascular response to early blockade of bradykinin receptor suggests that endogenous kinins play a role in the regulation of cardiovascular function in both sexes, but they may be functionally more important in the female rat.

Recognition of markers of response to potassium-canrenoate in essential hypertension.

Potassium canrenoate (K-Can) prevents hypertension in Milan hypertensive strain (MHS) but not in spontaneously hypertensive rats (SHR). Essential hypertensive patients (HT) may have differential sensitivity to diuretics, since a subgroup of HT insensitive to hydrochlorothiazide (HCTZ) but sensitive to K-Can has previously been found. The aims of this study were: 1) to seek markers of response in essential hypertensive patients selectively sensitive to K-Can: and 2) to test whether selective sensitivity to furosemide may also be demonstrated. After 2 weeks of placebo (P) 50 uncomplicated, mild to moderate HT (46 +/- 9 yrs, mean +/- SD) received K-Can (50 mg/day) for 4 weeks. After 2 more weeks of P, patients received HCTZ (25 mg) and furosemide (25 mg) for 4 weeks each in a single blind crossover design, with 2 weeks P between each treatment. Dosages were doubled after 2 weeks if diastolic blood pressure (DBP) was > 90 mmHg. Responders (R) were those HT whose DBP was < or = 90 mmHg and/or at least 10 mmHg lower than before treatment. Systolic blood pressure (SBP)/DBP was measured every 2 weeks with plasma renin activity (PRA), red blood cell Na(+)-K(+)-Cl- cotransport (COT) and Na(+)-K+ ATPase pump activity measured at the end of the first P period, and serum electrolytes at the end of each period. Four HT dropped out because of low compliance, 6 because of reversible side effects, and 1 because blood pressure was not back to pre-treatment value after the second placebo period.(ABSTRACT TRUNCATED AT 250 WORDS)

Chronic inhibition of bradykinin receptors alters cardiovascular function in rats with an excess of circulating vasoconstrictors.

1. The contribution of endogenous kinins to the regulation of blood pressure of angiotensin-treated rats was evaluated using the new bradykinin B2-receptor antagonist Hoe 140 (D-Arg[Hyp3,Thi5,D-Tic7,Oic8]-bradykinin). 2. Chronic intraperitoneal infusion of 20 nmol/day angiotensin II did not alter systolic blood pressure or plasma angiotensin II levels. A significant increase in plasma aldosterone and corticosterone levels was observed after 4 weeks (from 89 +/- 20 to 140 +/- 22 and from 147 +/- 30 to 225 +/- 33 pg/ml, respectively; P < 0.05). 3. Combined administration of 20 nmol/day angiotensin II and 75 nmol Hoe 140 induced a significant increase in systolic blood pressure from 126 +/- 3 to 142 +/- 3 and 137 +/- 3 mmHg, at 1 and 4 weeks, respectively (P < 0.05). This effect was not accompanied by significant changes in plasma angiotensin II concentration. The angiotensin-induced increase in plasma levels of aldosterone and corticosterone was not altered by the antagonist Hoe 140. 4. These findings indicate that blockade of endogenous kinin receptors enhances the slow pressor effect induced by angiotensin II. Therefore, endogenous kinins may play a role in preventing the cardiovascular effects of an excess of vasoconstrictors.

Kinin antagonist blunts the diuretic effect of furosemide in deoxycorticosterone-treated rats.

Furosemide at a dose of 3 mg/kg body wt increased urinary volume (vehicle: 12.8 +/- 0.6; furosemide: 42.4 +/- 2.6 ml/8h, p < 0.01) and urinary sodium excretion (vehicle: 0.9 +/- 0.1; furosemide: 5.0 +/- 0.4 mM/8h, p < 0.01) in deoxycorticosterone-treated rats. These effects were associated to a decrease in mean blood pressure (from 122 +/- 4 to 113 +/- 3 mmHg, p < 0.01) and renal vascular resistances (from 15.6 +/- 0.6 to 14.3 +/- 0.7 RU, p < 0.05). The B2-receptor antagonist D-Arg[Hyp3,Thi5,D-Tic7,Oic8]-bradykinin significantly blunted the diuretic and natriuretic effect of furosemide and completely prevented the decrease in blood pressure and renal vascular resistances. The renal kallikrein-kinin system may modulate the diuretic and hemodynamic effects of furosemide in conditions of increased mineralcorticoid activity.

[Effect of chronic enalapril treatment on prorenin, renin and sex hormones during the menstrual cycle in patients with essential hypertension]. / Effetti del trattamento cronico con enalapril su prorenina, renina ed ormoni sessuali durante il ciclo mestruale in pazienti ipertese essenziali.

Since angiotensin II seems to be involved in the process of ovulation we studied the effect of chronic enalapril on plasma prorenin, renin, estradiol, progesterone, LH and FSH during the menstrual cycle in ten essential hypertensive women. Our data show that peripheral blockade of A I conversion does not affect the pituitary guidance and the ovarian hormonal response of the menstrual cycle and, we can speculate, that it does not interfere with the process of ovulation.

[Protective effect of a calcium antagonist against renal vasoconstriction induced by endothelin in the normotensive rat]. / Effetto protettivo di un calcio-antagonista nei confronti della vasocostrizione renale indotta dall'endotelina nel ratto normoteso.

We studied the effect of nifedipine, a dihydropyridine calcium antagonist, on the hemodynamic changes induced by endothelin, in awake normotensive rats. Endothelin (0.07-1.40 nmol/kg, e.v.) caused an initial hypotensive effect, followed by long lasting hypertension. Renal blood flow was reduced immediately and still remained below basal levels, at 30 minutes after endothelin injection. Nifedipine (1 mg/kg, i.p.) significantly prevented the effect of endothelin on mean blood pressure and induced a right-ward shift in the dose response curve of renal hemodynamic changes induced by endothelin. We conclude that treatment with calcium antagonist could be very useful in all those conditions in which systemic and regional vasocostriction is provoked by endothelin.

[Effect of menstrual cycle hormones on cation transport in the red-cell membrane]. / Influenze degli ormoni del ciclo mestruale sui trasporti cationici di membrana dei globuli rossi.

Erythrocyte cation transport, plasma prorenin and renin and sexual hormones were sequentially evaluated in 12 normal volunteers over the menstrual cycle. Na-K cotransport and Na-Li countertransport raised in 6 out of 12 subjects in synchronization with the ovulatory phase. When the maximal % variation (ovulatory phase) versus baseline (follicular phase) of the Na-K cotransport was plotted versus the maximal % increment of oestrogens. A direct, highly significant inverse correlation was observed (r = 0.904, p less than 0.001). Moreover, a highly significant inverse correlation between plasma prorenin and intraerythrocyte Na (r = -0.857, p less than 0.001) in the follicular phase was found. Our data suggest that erythrocyte cation transport can be influenced by sexual hormones in human.